Commonwealth Scientific and Industrial Research Organisation v BASF Plant Science GmbH - [2020] FCA 328

FEDERAL COURT OF AUSTRALIA

Commonwealth Scientific and Industrial Research Organisation v BASF Plant Science GmbH [2020] FCA 328

File number:

VID 42 of 2019

Judge:

BEACH J

Date of judgment:

12 March 2020

Catchwords:

PATENTS – production of long-chain polyunsaturated fatty acids – transgenic organisms – genes from unicellular algae coding for enzymes – appeal from Commissioner of Patents – application to amend specification – construction of specification – meaning of “substrate specificity” – stipulation of preference for conversion of alpha linolenic acid compared to linoleic acid – added matter – intermediate generalisation – s 102(1) of Patents Act 1990 (Cth) – whether specification as amended would claim or disclose matter that extends beyond specification as filed – relevance of analogous foreign law tests – s 76(2) of Patents Act 1977 (UK) – art 123(2) of European Patent Convention – comparison with s 102(1) test pre- Intellectual Property Laws Amendment (Raising the Bar) Act 2012 (Cth) – amendment impermissible – appeal allowed

Legislation:

Intellectual Property Laws Amendment (Raising the Bar) Act 2012 (Cth) Sch 1

Patents Act 1949 (UK) s 31

Patents Act 1977 (UK) s 76

Patents Act 1990 (Cth) ss 40, 102, 104

Patents Regulations 1991 (Cth) reg 10.2A

Cases cited:

AstraZeneca AB v Apotex Pty Ltd (2014) 226 FCR 324

Bonzel v Intervention Ltd (No 3) [1991] RPC 553

Buhler AG v FP Spomax SA [2008] FSR 27

Coopers Animal Health Australia Ltd v Western Stock Distributors Pty Ltd (1987) 15 FCR 382

Cytec Industries Inc v Nalco Company [2019] FCA 1800

European Central Bank v Document Security Systems Inc [2007] EWHC 600 (Pat)

Les Laboratoires Servier v Apotex Pty Ltd (2010) 273 ALR 630

LG Philips LCD Co Ltd v Tatung (UK) Ltd [2007] RPC 21

Lockwood Security Products Pty Ltd v Doric Products Pty Ltd (2004) 217 CLR 274

Meat and Livestock Australia Ltd v Cargill Inc (2018) 354 ALR 95

Meat & Livestock Australia Ltd v Cargill, Inc (No 2) (2019) 139 IPR 47

Napp Pharmaceutical Holdings Ltd v Ratiopharm GmbH [2009] RPC 18

Palmaz’s European Patents (UK) [1999] RPC 47

Palmaz’s European Patents (UK) [2000] RPC 631

Pilkin v Sony Australia Limited (No 2) [2019] FCA 980

RGC Mineral Sands Pty Ltd v Wimmera Industrial Minerals Pty Ltd (1998) 89 FCR 458

Nokia Corporation v IPCom GmbH & Co KG (No 3) [2013] RPC 5

Richardson-Vicks Inc’s Patent [1995] RPC 568

Smith & Nephew PLC v Convatec Technologies Inc [2013] RPC 8

Southco Inc v Dzus Fastener Europe Ltd [1990] RPC 587

Vector Corporation v Glatt Air Techniques Ltd [2008] RPC 10

Date of hearing:

20 and 21 February 2020

Registry:

Victoria

Division:

General Division

National Practice Area:

Intellectual Property

Sub-area:

Patents and associated Statutes

Category:

Catchwords

Number of paragraphs:

348

Counsel for the Appellant:

Mr BN Caine QC and Ms KB Beattie

Solicitor for the Appellant:

King & Wood Mallesons

Counsel for the Respondent:

Mr C Smith

Solicitor for the Respondent:

Watermark Intellectual Property Lawyers

ORDERS

VID 42 of 2019

BETWEEN:

COMMONWEALTH SCIENTIFIC AND INDUSTRIAL RESEARCH ORGANISATION

Appellant

AND:

BASF PLANT SCIENCE GMBH

Respondent

JUDGE:

BEACH J

DATE OF ORDER:

12 March 2020

THE COURT ORDERS THAT:

1.        The appeal be allowed.
2.        The decision of the Commissioner of Patents made on 10 December 2018 be set aside.
3.        Within 21 days of the date hereof the parties file and serve proposed minutes of orders and short submissions (limited to 3 pages) as to any necessary consequential orders including on the question of costs.

Note: Entry of orders is dealt with in Rule 39.32 of the Federal Court Rules 2011.

REASONS FOR JUDGMENT

BEACH J:

The Commonwealth Scientific and Industrial Research Organisation (CSIRO) appeals from a decision of a delegate of the Commissioner of Patents allowing amendments to Australian patent application 2013273704 entitled “Process for the production of polyunsaturated fatty acids in transgenic organisms” (the application) filed by BASF Plant Science GmbH (BASF).
The application relates to genes from a species of unicellular algae that code for enzymes which can be employed for the recombinant production of polyunsaturated fatty acids in plants. Broadly, it is directed to the production of long-chain polyunsaturated fatty acids in higher plants by incorporating genes that encode for relevant enzymes from microalgae, specifically one called Ostreococcus lucimarinus.
CSIRO contends that the amendments are not allowable because, as a result of the amendments, the amended specification would claim and disclose matter that extends beyond that disclosed in the complete specification as filed, and therefore are impermissible under s 102(1) of the Patents Act 1990 (Cth) (the Act).
The application is a divisional application of Australian patent application 2007304229 filed on 4 October 2007. But the application claims priority from European patent application 06121888.9 filed on 6 October 2006 (the priority date).
The application was filed on 19 December 2013. BASF requested three amendments to the application as filed prior to acceptance; those amendments were requested on 7 September 2015, 26 February 2016 and 26 April 2016.
The application was accepted with those amendments on 4 May 2016. The application as accepted was published on 19 May 2016. CSIRO filed a notice of opposition to the application as accepted on 19 August 2016.
On 7 June 2017, BASF filed a request to amend the specification of the application as accepted under s 104. That request was not allowed on the basis that, as a result of the proposed amendment, the specification would not comply with the requirements of s 40(3).
On 2 October 2017, BASF filed a further request to amend the specification of the application as accepted.
Details of these two requests were published in the Australian Official Journal of Patents on 30 November 2017. CSIRO filed a notice of opposition to any allowance of those amendments on 30 January 2018.
On 10 December 2018, a delegate of the Commissioner of Patents allowed the post-acceptance amendments sought. CSIRO has appealed that decision under s 104(7). The nature of the appeal before me is by way of a rehearing de-novo.
Now whether the proposed post-acceptance amendments are allowable is a question to be determined under s 102(1) of the Act as amended by the Intellectual Property Laws Amendment (Raising the Bar) Act 2012 (Cth) (the Raising the Bar Act). The amendments made to s 102(1) of the Act by item 29 of Sch 1 of the Raising the Bar Act apply in relation to amendments of complete specifications directed or requested to be made on or after 15 April 2013 if the amendments are in relation to complete patent applications made on or after that day (Raising the Bar Act, sch 1, item 55(9)(d)). As I have said, the application was filed on 19 December 2013.
Section 102(1) in its present and applicable form provides:
(1)       An amendment of a complete specification is not allowable if, as a result of the amendment, the specification would claim or disclose matter that extends beyond that disclosed in the following documents taken together:
(a)       the complete specification as filed;
(b)       other prescribed documents (if any).
For present purposes, the only “other prescribed document” (reg 10.2A of the Patents Regulations 1991 (Cth)) is the abstract that accompanied the application. But this is identical to page 1, lines 7 to 13 of the application as filed. Accordingly, consideration of the abstract adds nothing to the analysis.
Now it is not in doubt that the matter arising “as a result of the amendment” is identified by identifying the difference between the application as accepted and the application as proposed to be amended. One then compares what is disclosed in the application as proposed to be amended, of course read as a whole, with what is disclosed in the application as filed. If as a result of the amendment proposed, the specification would disclose or claim matter that extends beyond that disclosed in the application as filed, the amendment is not allowable. Moreover, part of the analysis concerning construing the application as filed is to be undertaken through the eyes of a person skilled in the art at the relevant time.
On the present appeal CSIRO filed and served expert evidence from Dr Surinder Singh who was at the time of preparing his written evidence a Group Leader and Chief Research Scientist in CSIRO’s Agriculture and Food Unit. BASF filed and served expert evidence from Dr David Stalker. The experts met in conclave and subsequently produced a joint expert report. I took their evidence in a concurrent evidence session with the debate between them largely focused upon the question of substrate specificity concerning polyunsaturated fatty acids that was addressed in the application as filed.
As I have indicated, the application relates to the field of isolating polynucleotides which code for enzymes that can be used in the biosynthetic pathway of long-chain polyunsaturated fatty acid synthesis and introducing these polynucleotides into plants (e.g. oil seed crops) by genetic engineering with the intention that long-chain polyunsaturated fatty acids will be produced in such plants. The work in this field is predominantly done by senior research scientists with significant research experience in relation to the synthesis of fatty acids in transgenic plants. In the present case, the hypothetical person skilled in the art at the relevant time is such a scientist involved in metabolic/genetic engineering of plants to produce fatty acids.
Dr Singh has a PhD from the University of Adelaide in plant physiology and biochemistry. He has extensive experience in metabolic/genetic engineering of plants for the development of plant oils, including fatty acids. He is a named author of 102 peer-reviewed articles and papers, including 75 publications directed to plant lipids and modifying plant oils. He is a person skilled in the field of the application.
Dr Stalker also holds a PhD from the University of Cincinnati in the field of cell biology and bio-chemistry. He has worked as a scientist in the field of molecular biology for over 45 years. For a substantial part of that period he has researched and commercialised innovations in plant biotechnology, including creating genetically engineered plants. He has also published a substantial number of peer-reviewed articles and papers in this field. He has been a professor of biotechnology at RMIT and is the founder and a director of Generic Genetics Group Pty. Ltd. He is similarly skilled in the relevant field.
For the reasons that follow, I am clearly satisfied that the proposed amendments are impermissible and that the appeal should be allowed.
Let me begin with some science before turning to discuss and compare the application as filed, the application as accepted and the application as proposed to be amended.

SOME SCIENCE

Let me begin by setting out a functional glossary, that is, functional to following my reasons and functional to appreciating the context of the invention the subject of the application, of the following terms:
•         Alpha(α)-linolenic acid
The starting material for the synthesis of omega(ω)-3 long-chain polyunsaturated fatty acids.
•         CoA-dependent enzyme
An enzyme that predominantly acts on acyl-Coenzyme A (CoA) bound fatty acid substrates in the cytosolic acyl-CoA pool as opposed to acting on phosphatidylcholine bound fatty acid substrates in the endoplasmic reticulum-associated phospholipid pool.
•         Desaturase
An enzyme that catalyses the insertion of a carbon to carbon double bond in the carbon backbone by removing two hydrogen atoms from a fatty acid.
•         Elongase
An enzyme that catalyses the addition of two carbon atoms to the terminal carboxyl end of the fatty acid, extending the length of a fatty acid.
•         Enzyme
A protein molecule that functions as a catalyst for biochemical reactions.
•         Fatty acid
A carboxylic acid consisting of a chain of carbon atoms bonded to hydrogen atoms (forming a carbon backbone) having a carboxyl group (COOH) at one end and a methyl group (CH₃) at the other end.
•         Linoleic acid
The starting material for the synthesis of omega(ω)-6 long-chain polyunsaturated fatty acids, for example arachidonic acid.
•         Long chain PUFA (LC-PUFA)
A fatty acid having a carbon backbone that is at least 20 carbon atoms long and contains two or more double bonds.
•         Ostreococcus lucimarinus (O. lucimarinus)
A unicellular algae.
•         Omega(ω)-3 fatty acid
A fatty acid with the last carbon to carbon double bond three carbon atoms away from the terminal methyl group, for example, eicosapentaenoic acid and docosahexaenoic acid.
•         Omega(ω)-6 fatty acid
A fatty acid with the last carbon to carbon double bond six carbons atoms away from the terminal methyl group.
•         Polynucleotide
A nucleic acid sequence made up of nucleotides that code for genetic information.
•         Polypeptide
A sequence of amino acids forming a protein.
•         Polyunsaturated fatty acids (PUFAs)
Fatty acids having two or more carbon to carbon double bonds in the fatty acid chain. PUFAs are categorised by the number of carbon atoms in the chain, the number of unsaturated (double) bonds, and the position of those double bonds. PUFAs can be classified into two main groups, namely, omega(ω)-3 fatty acids and omega(ω)-6 fatty acids.
•         Recombinant production/recombinant methods
A combination of genetic material (DNA) from different organisms.
•         Saturated fatty acid
A fatty acid containing no carbon to carbon double bonds.
•         Specificity
For the moment I will define this to be the ability of an enzyme to discriminate between competing potential substrates.
•         Substrate
The molecule upon which an enzyme acts.
•         Unsaturated fatty acid
A fatty acid having at least one carbon to carbon double bond, rather than only single bonds.
It is also necessary to set out the following list of the fatty acids referenced in the relevant biosynthetic pathway that I will discuss later:

List of fatty acids referenced in the biosynthetic pathway

Abbreviation

Carbons

(No of carbons: no of double bonds)

Name

OA

18:1

Oleic acid

LA

18:2

Linoleic acid

ALA

18:3

Alpha-linolenic acid

GLA

18:3

Gamma-linolenic acid

SDA

18:4

Stearidonic acid

EDA

20:2

Eicosadienoic acid

ETrA

20:3

Eicosatrienoic acid

DGLA

20:3

Dihomo-gamma-linolenic acid

ETA

20:4

Eicosatetraenoic acid

AA (or ARA)

20:4

Arachidonic acid

EPA

20:5

Eicosapentaenoic acid

DPA

22:5

Docosapentaenoic acid

DHA

22:6

Docosahexaenoic acid

(a) Enzymes

Let me begin my discussion of some of the science by talking about enzyme function. What I have set out below on this topic and later biochemical topics is a composite of the evidence from the two experts that I did not understand to be in issue concerning the applicable common general knowledge relevant to the task of construing the application as filed, accepting of course that the relevant legal questions are for me as to what it discloses.
In a biological setting, enzymes are typically proteins which function as catalysts for biochemical reactions. An enzyme works on a molecule known as the substrate, and the molecules formed by the reaction are known as the products.
A protein is made up of one or more chains of amino acids. A protein will be an enzyme if it performs the role of catalysing a biochemical reaction. The chemical structure that a protein enzyme has and its function is dictated by the particular order or sequence of the amino acids that make up that enzyme. Changing that sequence of amino acids may create a different protein enzyme, which may or may not then have a different function.
The sequence of amino acids that form a protein enzyme is in turn dictated by the gene sequence which encodes the protein enzyme. A gene sequence is a chain of nucleotide bases. The information in a gene sequence is stored as a “code” utilising four nucleotides, albeit in triplets of three nucleotides known as codons. The four nucleotides are adenine (A), cytosine (C), guanine (G) and thymine (T). The sequence of these nucleotides determines the information available for building and maintaining an organism. I have discussed the relevant concepts elsewhere (Meat and Livestock Australia Ltd v Cargill Inc (2018) 354 ALR 95 (Meat and Livestock (No 1)) at [21] to [66]). Judicial notice can be taken of the fact that such concepts have been well known to biochemists, biologists and experts in derivative fields for some decades.
Whilst a particular gene sequence may encode a specific protein enzyme, it is possible that a different gene sequence may still encode the same enzyme. So, the same protein enzyme could be encoded by two or more different gene sequences. Different codons can code for the same amino acid, for example, TTT and TTC both code for phenylalanine. So, variation at the DNA level does not necessarily produce a protein having altered structure and function.
On the other hand, even a single nucleotide change to a DNA sequence that results in a single amino acid change at a given point can change the structure of the encoded protein enzyme and therefore alter its function. For example, a single nucleotide change such as TTT to TAT produces a different amino acid that has different biochemical properties; in this example the change is from phenylalanine (non-polar) to tyrosine (polar).
Now an enzyme works on the substrate. An enzyme may bind to and convert a single substrate only. Alternatively, an enzyme may bind to and convert multiple substrates. An enzyme may also catalyse more than one reaction. For example, an enzyme may only act on molecules that have specific functional groups, for example amino, phosphate or methyl groups, or on a particular type of chemical bond, or on a particular steric or optical isomer.
Generally speaking, the “substrate specificity” of an enzyme refers to the ability of that enzyme to discriminate among competing potential substrates and to bind to a specific substrate or substrates. I will return to this issue and discuss it in more detail later. Substrate specificity is the result of structural and conformational complementarity between the enzyme and the relevant substrate or substrates. Where an enzyme can bind to more than one substrate, the enzyme may show a preference for binding to one substrate over another at differing rates or even absolutely.
Whether an enzyme has specificity for one or more relevant substrates or is able to effectively perform one or more catalytic reactions will be determined by the amino acid sequence of the enzyme. Consequently, variation in amino acid sequence may influence whether an enzyme might bind to one or multiple substrates or is able to perform one or more catalytic reactions. Sequence variation may also influence an enzyme’s affinity for one or more substrates, therefore impacting substrate specificity for one or more substrates and the enzyme’s preference when able to bind to more than one substrate.
Some enzymes can catalyse more than one type of reaction. An enzyme that catalyses two distinct reactions is known as bifunctional. Further, the substrate specificity for the enzyme may depend on the relevant function or catalytic activity. Further, the substrate specificity for one catalytic activity may be different from the substrate specificity for the other catalytic function. In other words, substrate specificity may need to be considered in the context of a relevant function or catalytic activity.
The efficiency of the enzyme to convert a substrate or substrates into a relevant product or products refers to the rate at which the substrate is converted into the product. Efficiency can be dependent on amino acid sequence. A change in sequence could alter efficiency by altering substrate specificity, the strength at which the enzyme binds to the substrate being its binding affinity or the rate of catalysis.
Enzymes that perform the same or a similar function may have a degree of sequence relatedness, both in terms of the sequence of amino acids that make up each enzyme, and in terms of the DNA sequence that encodes for each enzyme.

(b) Polyunsaturated fatty acids

Let me now say something about fatty acids and particularly polyunsaturated fatty acids (PUFAs) and long-chain polyunsaturated fatty acids (LC-PUFAs).
A fatty acid is the basic building block of fats and lipids in plants, animals, microorganisms and food. A fatty acid is a carboxylic acid consisting of a hydrocarbon chain that forms a carbon backbone. At one end of the carbon backbone is a carboxyl group (COOH), which is called the terminal carboxyl end, and at the other end of the chain is a methyl group (CH₃), which is called the terminal methyl end. Fatty acids are either saturated, that is, the carbon backbone contains no double bonds between the carbon atoms, or unsaturated, that is, the carbon backbone contains one or more carbon to carbon double bonds (C=C double bonds). Monounsaturated fatty acids contain one C=C double bond, and PUFAs are fatty acids that contain more than one C=C double bond.
Fatty acids differ in the length of the carbon backbone, and can be categorised from short chain fatty acids to very long chain fatty acids. They contain an even number of carbons in the carbon backbone. LC-PUFAs are fatty acids that contain a carbon backbone that is at least 20 carbons long and contains multiple C=C double bonds.
PUFAs can be characterised as either omega-3 (ω3) or omega-6 (ω6) fatty acids based on the position of the first C=C double bond from the terminal methyl group. In particular, ω3 fatty acids have the last C=C double bond 3 carbon atoms away from the terminal methyl group whereas ω6 fatty acids have the last C=C double bond 6 carbon atoms away from the terminal methyl group.

LC-PUFAs have important roles in human health and development, with many studies predating the priority date indicating that deficiencies in these fatty acids can increase, for example, the risk or severity of cardiovascular disease, inflammatory diseases, and neuropsychiatric disorders such as depression or dementia.
Nutritionally important ω6 LC-PUFAs include arachidonic acid (AA) and nutritionally important ω3 LC-PUFAs include eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). These fatty acids are either directly available as components of the diet or produced from the two essential ω6- and ω3- fatty acids, linoleic acid (LA) and α-linolenic acid (ALA). Although humans can synthesize both EPA and DHA from ALA, direct uptake of EPA and DHA from the diet, for example from marine fish (fish oils), is a significantly more important source of ω3 LC-PUFAs for humans.
As a result of a plateau and potentially a decline in marine fish stocks since the early 1990s, significant research effort has focused on producing ω3 LC-PUFAs in alternative sources including aquaculture, cultured microalgae and oilseed crops that have been engineered to produce ω3 LC-PUFAs, such as EPA and DHA. Because of the production capacity and relatively low production cost of PUFAs from plants, oilseed crops that had been genetically engineered to synthesise fatty acids were viewed as a significant goal as a sustainable alternative source of ω3 LC-PUFAs.
Before the priority date, transgenic plants had already been produced which synthesized EPA, docosapentaenoic acid (DPA) and DHA in their seed.
Prior to the priority date, researchers had been experimenting with introducing foreign genes into plants to alter biological properties and/or to add or enhance physical attributes, such as resistance to stress and disease or herbicide resistance. A plant that is modified using genetic techniques is usually referred to as a genetically engineered plant or a transgenic plant.
In a transgenic plant, the source of any introduced foreign gene is relevant insofar as the DNA or amino acid sequence determines the encoded protein’s structure and/or function, such that the introduction of a foreign gene encoding an enzyme from one source can have a different impact in the transgenic plant compared to introduction of a foreign gene encoding an enzyme of a different source, even where both enzymes have the same or similar function.

(c) Synthesising long-chain polyunsaturated fatty acids

Let me elaborate further on LC-PUFA plant metabolic engineering.
Prior to the priority date research groups were working on isolating and cloning genes from various fungi, protists and microalgae that encode each of the desaturases and elongases involved in the aerobic pathway for LC-PUFA synthesis. The availability of genes that encode the aerobic LC-PUFA biosynthetic enzymes led to attempts to introduce these genes into plants (such as oil seed crops) via recombinant methods and to express these enzymes in these plants to produce LC-PUFAs. Recombinant methods of course combine DNA sequences from different organisms, creating a new recombinant DNA molecule that is not otherwise found in nature.
As I have indicated, fatty acids form an important component of soluble fats in living cells and consist of a chain of an even number of carbon atoms, with hydrogen atoms along the length of the chain. At one end of the chain is a methyl group. A carboxyl group is at the other end.
And as I have said, PUFA is a term used to describe fatty acid chains that contain two or more carbon-carbon double bonds. In plants, PUFAs occur predominantly as chains having 16, 18, 20 or 22 carbons, hence are referred to as C16-, C18-, C20- and C22-PUFAs, respectively.
LC-PUFAs (20 carbons or longer) such as EPA and DHA play important roles in human health and are intrinsic to diet.
As I have said, humans can synthesise LC-PUFAs through dietary intake of precursor fatty acids. Humans can also obtain LC-PUFAs via direct dietary intake, e.g. by consuming fish and fish-derived oils that contain EPA and DHA. However, poor diets combined with decreasing global fish stocks have raised the importance of obtaining LC-PUFAs from other sources.
Now whilst higher plants generally lack the metabolic pathway for producing fatty acids longer than 18 carbons in length, since the late 1990s research has intensified in the field of transgenic oilseed plants. Transgenic oilseed plants are plants that are sources of short-chain fatty acids which are modified using genetic engineering techniques to introduce foreign genes to enable the production of LC-PUFAs like EPA and DHA. Genes encoding enzymes that enable LC‑PUFA production can be sourced from fungi, microalgae and lower plants which are known to produce LC-PUFAs.
Let me delve further into the detail and now say something about ω3 LC-PUFA synthesis in plants.
All higher plants have the enzymatic complex to synthesize the PUFAs that contain a carbon chain that is 18 carbons long, for example, LA and ALA, but do not possess the enzymes to convert these PUFAs into LC-PUFAs, that is, containing a chain of at least 20 carbon atoms long. The synthesis of LC-PUFAs in higher plants therefore requires the introduction of genes that encode all of the biosynthetic enzymes required to convert either LA into ω6 LC-PUFAs such as AA, or to convert ALA into ω3 LC-PUFAs such as EPA and DHA. These genes can be obtained from a range of organisms that synthesise LC-PUFAs, such as marine bacteria, fungi, protists and microalgae. LC-PUFA synthesis in these organisms involves either aerobic or anaerobic pathways.
The aerobic pathway consists of a series of distinct desaturation steps, that is, for each such step the insertion of a C=C double bond into the carbon backbone, and elongation steps, that is, for each such step the insertion of two carbons to the terminal carboxyl group of the fatty acid. Such steps will convert either LA into LC-PUFAs such as AA, EPA and DHA or convert ALA into LC-PUFAs such as EPA and DHA. This conversion of LA or ALA into these LC-PUFAs is shown schematically as follows:

Schematic representation of the conversion of LA or ALA into LC-PUFAs

Each of the fatty acids listed in the figure is referred to by its carbon chain length (18, 20 or 22) and the number of C=C double bonds (2, 3, 4, 5 or 6). For instance, LA is referred to as a C18:2 PUFA indicating that it has 18 carbon atoms in its carbon chain and 2 C=C double bonds. Further, the application as filed often includes a reference to whether the PUFA is an omega-6 (ω6) or omega-3 (ω3) PUFA and which carbon atoms along the chain include the C=C double bonds. For example, LA is denoted as a ω6-C18:2^Δ9,12 fatty acid, where “Δ9,12” indicates that the 2 C=C double bonds are located at the ninth and twelfth carbon atoms, counting the carbon atoms from the terminal carboxyl end rather than the terminal methyl group. In fatty acid biosynthesis, the “delta” symbol (Δ) indicates the position in the fatty acid at which an enzyme creates a double bond.
In the above schematic, the desaturation steps are undertaken by enzymes known as desaturases, and the elongation steps are undertaken by enzymes known as elongases.
In the above schematic, the first committed step in the ω6 and ω3 pathways is desaturation of LA and ALA, respectively, by a Δ6-desaturase which introduces a C=C double bond between carbons 6 and 7 of LA and ALA to produce GLA or SDA, respectively.
The C18 fatty acids, GLA and SDA, are then elongated by two carbons by a Δ6-elongase to produce the LC-PUFA, di-homo-γ-linolenic acid (DGLA) and eicosatetraenoic acid (ETA), respectively.
The products of the Δ6-elongation, DGLA and ETA, may then be desaturated by a Δ5-desaturase which introduces a C=C double bond between carbons 5 and 6 of DGLA and ETA to produce AA and EPA, respectively. This pathway, that is, a Δ6-desaturase followed by a Δ6-elongase and then a Δ5-desaturase, is often referred to as the Δ6-desaturase pathway.
An alternative aerobic pathway, often referred to as the alternate Δ9-elongase/Δ8-desaturase pathway, starts with elongation by a Δ9-elongase which adds two carbons to LA and ALA, followed by two successive desaturation reactions. The first desaturation reaction is catalysed by a Δ8-desaturase which introduces a double bond between carbons 8 and 9, producing DGLA and ETA, and the second is catalysed by a Δ5-desaturase, producing AA and EPA. The Δ9-elongase/ Δ8-desaturase pathway is also depicted in the above schematic on the far left hand side and far right hand side, producing the intermediates eicosadienoic acid (EDA) and eicosatrienoic acid (ETrA).
In both pathways, EPA is subsequently elongated by two carbons by a Δ5-elongase to DPA. And then DPA may be desaturated by a Δ4-desaturase which introduces a double bond between carbons 4 and 5 to yield DHA.
As the above schematic indicates, the ω6 PUFAs include LA and fatty acids derived from LA by desaturation and elongation. The ω6 PUFAs include GLA, 18:3^Δ6,9,12, EDA, 20:2^Δ11,14, DGLA, 20:3^Δ8,11,14 and AA, 20:4^Δ5,8,11,14.
The ω3 fatty acids include ALA and fatty acids derived from ALA by desaturation and elongation. The ω3 PUFAs include SDA, 18:4^Δ6,9,12,15, ETrA, 20:3^Δ11,14,17, ETA, 20:4^Δ8,11,14,17, EPA, 20:5^{Δ5,8,11,14,17}, DPA, 22:5^{Δ7,10,13,16,19} and DHA, 22:6^{Δ4,7,10,13,16,19}.
I should make two other points at this stage.
First, in terms of driving fatty acid biosynthesis, the particular structure of the enzyme that is used in the pathway can have an influence over enzyme functionality and efficiency. For example, variation in the amino acid sequence encoding a Δ6-desaturase may influence the enzyme’s ability to bind to one or both of LA and ALA, as well as the efficiency of conversion to GLA and SDA respectively.
Second, a Δ6-desaturase in the context of the above schematic will always utilise LA or ALA or both as substrates to introduce a double bond between carbons 6 and 7 from the carboxyl end of the acyl groups of LA and/or ALA. But an enzyme that has Δ6-desaturase catalytic activity may also have other catalytic activity.

(d) A bit more on substrate specificity and catalysis

I said that I would return to say something more on the science concerning substrate specificity. Let me do so now.
In evidence before me were only incomplete extracts from Lehninger AL’s Biochemistry (2^nd edition, 1975, Worth Publishers Inc (N.Y.)) and Lehninger: Principles of Biochemistry edited by Nelson DL and Cox MM (4^th edition, 2005, W.H Freeman and Company) concerning enzymes including their structure and catalytic function. The extracts tendered in part dealt with substrate specificity. It was necessary for me to procure with the assistance of the Court’s Victorian librarian, Mr Michael Coats, other parts of Chapter 6 of the 4^th edition in order to put into context the extract that had been given to me. For what it is worth, I was also able to procure and confirm that the current 7^th edition (2017) did not substantially differ from the 4^th edition as to the matters that I am concerned with; in any event it is well after the relevant date applicable to the construction questions. The following may be distilled from the 2^nd and 4^th editions, with the 7^th edition put to one side.
First, generally speaking, one of the notable features of enzymes is their specificity of action when compared with synthetic catalysts or inorganic catalysts. And but for such specificity, one would be swamped with unwanted if not deleterious side reactions and collateral products.
Second, unlike most other catalysts, enzymes have very significant catalytic power in dilute aqueous solutions at moderate temperatures and in biological pH ranges.
Third, enzymes vary in their specificity. Some will have near absolute specificity for a particular substrate. Others can bind to a whole class of molecules being structurally related substrates, but at different rates. Moreover, as the 2^nd edition explained, “substrate molecules generally reflect, by the principle of complementarity, the structure of the active site of the enzyme in two distinctive structural features: (1) the substrate must have a susceptible chemical bond that can be attacked by the enzyme; (2) it usually has some other structural feature required for its binding to the enzyme active site, presumably to position the substrate molecule in the proper geometrical relationship so that the susceptible bond can be attacked.”
Clearly, in the context of the 2^nd edition extract that I was given, specificity relates to the question of the enzyme bonding to the substrate. In this context, it is concerned with the ability of the enzyme to discriminate between a substrate and a competing molecule. Let me elaborate further by reference to section 6.2 of the 4^th edition and distinguish between the concepts of substrate specificity and catalytic effect, albeit of course related. I have done so because it was not entirely clear to me during the concurrent evidence session that the experts had sufficiently described the relevant concepts to my satisfaction; this of course reveals my own limitations rather than those of the experts. At one stage there appeared to be a blend between specificity and catalytic function and mechanism. Moreover, at one stage it seemed to me that the concepts of substrate and product had also been blended, although I understood that a substrate transformed through intermediate steps to a product by one enzyme could then function as a substrate upon which another enzyme could act, so producing another product, and so on. Further, in terms of substrate specificity as a precursor to catalytic function, the type of bonding/binding was not entirely clear to me. So, it has become necessary to expand on some of these concepts by reference to one of the standard works in the area at the time. I would take the person skilled in the art to have been well aware of the following biochemical concepts when coming to read the specification at the relevant time.
In terms of substrates, it is explained:
The distinguishing feature of an enzyme-catalyzed reaction is that it takes place within the confines of a pocket on the enzyme called the active site. The molecule that is bound in the active site and acted upon by the enzyme is called the substrate. The surface of the active site is lined with amino acid residues with substituent groups that bind the substrate and catalyze its chemical transformation. Often, the active site encloses a substrate, sequestering it completely from solution. The enzyme-substrate complex, whose existence was first proposed by Charles-Adolphe Wurtz in 1880, is central to the action of enzymes. It is also the starting point for mathematical treatments that define the kinetic behaviour of enzyme-catalyzed reactions and for theoretical descriptions of enzyme mechanisms (emphasis in original.)
The mechanics of catalysis are then explained in the following terms:
A simple enzymatic reaction might be written
E + S ⇌ ES ⇌ EP ⇌ E + P
where E, S, and P represent the enzyme, substrate, and product; ES and EP are transient complexes of the enzyme with the substrate and with the product.
The function of a catalyst is to increase the rate of a reaction. Catalysts do not affect reaction equilibria. Any reaction, such as S ⇌ P, can be described by a reaction coordinate diagram (Fig. 6-2), a picture of the energy changes during the reaction. Energy in biological systems is described in terms of free energy, G. In the coordinate diagram, the free energy of the system is plotted against the progress of the reaction (the reaction coordinate). The starting point for either the forward or the reverse reaction is called the ground state, the contribution to the free energy of the system by an average molecule (S or P) under a given set of conditions. To describe the free-energy changes for reactions, chemists define a standard set of conditions (temperature 298 K; partial pressure of each gas 1 atm, or 101.3 kPa; concentration of each solute 1 M) and express the free-energy change for this reacting system as ΔG, the standard free-energy change. Because biochemical systems commonly involve H⁺ concentrations far below 1 M, biochemists define a biochemical standard free-energy change, ΔG’, the standard free-energy change at pH 7.0.
FIGURE 6-2 Reaction coordinate diagram for a chemical reaction.
The free energy of the system is plotted against the progress of the reaction S P. A diagram of this kind is a description of the energy changes during the reaction, and the horizontal axis (reaction coordinate) reflects the progressive chemical changes (e.g., bond breakage or formation) as S is converted to P. The activation energies, ΔG^±, for the S P reactions are indicated ΔG’ is the overall standard free-energy change in the direction S P.
The equilibrium between S and P reflects the difference in the free energies of their ground states. In the example shown in Figure 6-2, the free energy of the ground state of P is lower than that of S, so ΔG’ for the reaction is negative and the equilibrium favors P. The position and direction of equilibrium are not affected by any catalyst.
A favorable equilibrium does not mean that the S P conversion will occur at a detectable rate. The rate of a reaction is dependent on an entirely different parameter. There is an energy barrier between S and P: the energy required for alignment of reacting groups, formation of transient unstable charges, bond rearrangements, and other transformations required for the reaction to proceed in either direction. This is illustrated by the energy “hill” in Figure 6-2. To undergo reaction, the molecules must overcome this barrier and therefore must be raised to a higher energy level. At the top of the energy hill is a point at which decay to the S or P state is equally probable (it is downhill either way). This is called the transition state. The transition state is not a chemical species with any significant stability and should not be confused with a reaction intermediate (such as ES or EP). It is simply a fleeting molecular moment in which events such as bond breakage, bond formation, and charge development have proceeded to the precise point at which decay to either substrate or product is equally likely. The difference between the energy levels of the ground state and the transition state is the activation energy, ΔG^±. The rate of a reaction reflects this activation energy: a higher activation energy corresponds to a slower reaction. Reaction rates can be increased by raising the temperature, thereby increasing the number of molecules with sufficient energy to overcome the energy barrier. Alternatively, the activation energy can be lowered by adding a catalyst. Catalysts enhance reaction rates by lowering activation energies (emphasis in original).

Further, it is explained that any reaction may have several steps, involving the formation and decay of transient chemical species called reaction intermediates. A reaction intermediate is any species on the reaction pathway that has a finite chemical lifetime. When the S ⇌ P reaction is catalyzed by an enzyme, the ES and EP complexes can be considered intermediates, even though S and P are stable chemical species. Further, when several steps occur in a reaction, the overall rate is determined by the step (or steps) with the highest activation energy; this is called the rate-limiting step.
The question was then posed as to what was the source of the energy for the dramatic lowering of the activation energies for specific reactions. The answer to the question had two distinct but related parts and was given in the following terms:
The first lies in the rearrangements of covalent bonds during an enzyme-catalyzed reaction. Chemical reactions of many types take place between substrates and enzymes’ functional groups (specific amino acid side chains, metal ions, and coenzymes). Catalytic functional groups on an enzyme may form a transient covalent bond with a substrate and activate it for reaction, or a group may be transiently transferred from the substrate to the enzyme. In many cases, these reactions occur only in the enzyme active site. Covalent interactions between enzymes and substrates lower the activation energy (and thereby accelerate the reaction) by providing an alternative, lower-energy reaction path.
The second part of the explanation lies in the non-covalent interactions between enzyme and substrate. Much of the energy required to lower activation energies is derived from weak, noncovalent interactions between substrate and enzyme. What really sets enzymes apart from most other catalysts is the formation of a specific ES complex. The interaction between substrate and enzyme in this complex is mediated by the same forces that stabilize protein structure, including hydrogen bonds and hydrophobic and ionic interactions. Formation of each weak interaction in the ES complex is accompanied by release of a small amount of free energy that provides a degree of stability to the interaction. The energy derived from enzyme-substrate interaction is called binding energy, ΔG_B. Its significance extends beyond a simple stabilization of the enzyme-substrate interaction. Binding energy is a major source of free energy used by enzymes to lower the activation energies of reactions (emphasis in original.)
Two fundamental and interrelated principles provide a general explanation for how enzymes use noncovalent binding energy:
1. Much of the catalytic power of enzymes is ultimately derived from the free energy released in forming many weak bonds and interactions between an enzyme and its substrate. This binding energy contributes to specificity as well as to catalysis.
2. Weak interactions are optimized in the reaction transition state; enzyme active sites are complementary not to the substrates per se but to the transition states through which substrates pass as they are converted to products during an enzymatic reaction.
Later it was said:
The same binding energy that provides energy for catalysis also gives an enzyme its specificity, the ability to discriminate between a substrate and a competing molecule. Conceptually, specificity is easy to distinguish from catalysis, but this distinction is much more difficult to make experimentally, because catalysis and specificity arise from the same phenomenon. If an enzyme active site has functional groups arranged optimally to form a variety of weak interactions with a particular substrate in the transition state, the enzyme will not be able to interact to the same degree with any other molecule. For example, if the substrate has a hydroxyl group that forms a hydrogen bond with a specific Glu residue on the enzyme, any molecule lacking a hydroxyl group at that particular position will be a poorer substrate for the enzyme. In addition, any molecule with an extra functional group for which the enzyme has no pocket or binding site is likely to be excluded from the enzyme. In general, specificity is derived from the formation of many weak interactions between the enzyme and its specific substrate molecule. (my emphasis.)
So, the mechanism for catalysis is separate from although of course related to the question of specificity. Moreover, as the authors explain, it is the weak interactions between the enzyme and the substrate that provide a substantial driving force for catalysis. And it was explained:
The groups on the substrate that are involved in these weak interactions can be at some distance from the bonds that are broken or changed. The weak interactions formed only in the transition state are those that make the primary contribution to catalysis. The requirement for multiple weak interactions to drive catalysis is one reason why enzymes (and some coenzymes) are so large. An enzyme must provide functional groups for ionic, hydrogen-bond, and other interactions, and also must precisely position these groups so that binding energy is optimized in the transition state. Adequate binding is accomplished most readily by positioning a substrate in a cavity (the active site) where it is effectively removed from water. The size of proteins reflects the need for superstructure to keep interacting groups properly positioned and to keep the cavity from collapsing.

THE EVOLUTION OF THE APPLICATION

At this point let me go through the evolution of the application, dealing first with the application as filed, then the application as accepted, and then the proposed amendments to the application as accepted; such amendments in context will then need to be compared with the application as filed.

(a) Application as filed

The specification of the application as filed is entitled “Process for the production of polyunsaturated fatty acids in transgenic organisms”.
The specification commences (p 1 lines 5 to 13) with the following description of the field of the invention:
The present invention relates to polynucleotides from Ostreococcus lucimarinus which code for desaturases and elongases and which can be employed for the recombinant production of polyunsaturated fatty acids. The invention furthermore relates to vectors, host cells and transgenic nonhuman organisms which comprise the polynucleotides, and to the polypeptides encoded by the polynucleotides. Finally, the invention also relates to production processes for the polyunsaturated fatty acids and for oil, lipid and fatty acid compositions.
Ostreococcus lucimarinus (O. lucimarinus) is a species of unicellular algae.
Various health benefits of PUFAs (e.g. brain development and function; eye function; cholesterol levels; arthritis) are described (p 1, line 23 to p 3, line 4).
It is explained (p 1 lines 15 to 30):
Fatty acids and triacylglycerides have a multiplicity of applications in the food industry, in animal nutrition, in cosmetics and in the pharmacological sector. Depending on whether they are free saturated or unsaturated fatty acids or else triacylglycerides with an elevated content of saturated or unsaturated fatty acids, they are suitable for very different applications. Polyunsaturated fatty acids such as linoleic acid and linolenic acid are essential for mammals, since they cannot be produced by the latter. Polyunsaturated ω3-fatty acids and ω6-fatty acids are therefore an important constituent in animal and human nutrition.
Polyunsaturated long-chain ω3-fatty acids such as eicosapentaenoic acid (= EPA, C20:5^{Δ5,8,11,14,17}) or docosahexaenoic acid (= DHA, C22:6^{Δ4,7,10,13,16,19}) are important components in human nutrition owing to their various roles in health aspects, including the development of the child brain, the functionality of the eyes, the synthesis of hormones and other signal substances, and the prevention of cardiovascular disorders, cancer and diabetes (Poulos, A Lipids 30:1-14, 1995; Horrocks, LA and Yeo YK Pharmacol Res 40:211-225, 1999). This is why there is a demand for the production of polyunsaturated long-chain fatty acids.
At page 2, lines 8 to 19 the specification explains that LC-PUFAs, such as EPA, DHA, AA (ARA) and DPA are conventionally obtained from fish and not synthesised in oil producing crops. The following is said:
The various fatty acids and triglycerides are mainly obtained from microorganisms such as Mortierella and Schizochytrium or from oil-producing plants such as soybean, oilseed rape, algae such as Crypthecodinium or Phaeodactylum and others, where they are obtained, as a rule, in the form of their triacylglycerides (= triglycerides = triglycerols). However, they can also be obtained from animals, such as, for example, fish. The free fatty acids are advantageously prepared by hydrolysis. Very long-chain polyunsaturated fatty acids such as DHA, EPA, arachidonic acid (=ARA, C20:4^Δ5,8,11,14), dihomo-γ-linolenic acid (C20:3^Δ8,11,14) or docosapentaenoic acid (DPA, C22:5^{Δ7,10,13,16,19}) are not synthesized in oil crops such as oilseed rape, soybean, sunflower or safflower. Conventional natural sources of these fatty acids are fish such as herring, salmon, sardine, redfish, eel, carp, trout, halibut, mackerel, zander or tuna, or algae.
The specification acknowledges that, “there has been no lack of attempts in the past to make available genes which are involved in the synthesis of fatty acids or triglycerides for the production of oils in various organisms …” (p 3 lines 5 to 7). Such prior art is then described (p 3, line 8 to p 4, line 26).
It is then explained (p 4 line 27 to p 5 line 12):
Depending on their desaturation pattern, the polyunsaturated fatty acids can be divided into two large classes, viz. ω6- or ω3-fatty acids, which differ with regard to their metabolic and functional activities.
The starting material for the ω6-metabolic pathway is the fatty acid linoleic acid (18:2^Δ9,12) while the ω3-pathway proceeds via linolenic acid (18:3^Δ9,12,15). Linolenic acid is formed by the activity of an ω3-desaturase (Tocher et al. 1998, Prog. Lipid Res. 37, 73-117; Domergue et al. 2002, Eur. J. Biochem. 269, 4105-4113).
Mammals, and thus also humans, have no corresponding desaturase activity (Δ12- and ω3-desaturase) and must take up these fatty acids (essential fatty acids) via the food. Starting with these precursors, the physiologically important polyunsaturated fatty acids arachidonic acid (= ARA, 20:4^Δ5,8,11,14), an ω6-fatty acid and the two ω3-fatty acids eicosapentaenoic acid (= EPA, 20:5^{Δ5,8,11,14,17}) and docosahexaenoic acid (DHA, 22:6^{Δ4·7,10,13,17,19}) are synthesized via the sequence of desaturase and elongase reactions. The application of ω3-fatty acids shows the therapeutic activity described above in the treatment of cardiovascular diseases (Shimikawa 2001, World Rev. Nutr. Diet. 88, 100-108), inflammations (Calder 2002, Proc. Nutr. Soc. 61, 345-358) and arthritis (Cleland and James 2000, J. Rheumatol. 27, 2305-2307).
The specification identifies that it would be advantageous to introduce genes that encode for the enzymes for LC-PUFA biosynthesis in higher plants (p 6 lines 9 to 16):
To this end, it is advantageous to introduce, into oil crops, genes which encode enzymes of the LCPUFA biosynthesis via recombinant methods and to express them therein. These genes encode for example Δ6-desaturases, Δ6-elongases, Δ5‑desaturases or Δ4-desaturases. These genes can advantageously be isolated from microorganisms and lower plants which produce LCPUFAs and incorporate them in the membranes or triacylglycerides. Thus, it has already been possible to isolate Δ6‑desaturase genes from the moss Physcomitrella patens and Δ6-elongase genes from P. patens and from the nematode C. elegans.
The specification acknowledges that such plants, comprising and expressing genes encoding the enzymes necessary for LC-PUFA biosynthesis, have been described in the prior art (p 6 lines 18 to 22), but that they produce LC-PUFAs in “amounts which require further optimization for processing the oils which are present in the plants”.
The object of the invention is then stated (p 6, lines 24 to 30):
To make possible the fortification of food and of feed with these polyunsaturated fatty acids, there is therefore a great need for means and measures for a simple inexpensive production of these polyunsaturated fatty acids, specifically in eukaryotic systems. The object of the present invention would therefore be the provision of such means and measures. This object is achieved by the use forms which are described in the patent claims and hereinbelow.
The invention the subject of the application is then discussed (p 6 line 31 to p 7 line 2):
The invention, the subject of the present application, is directed to the following:
a CoA-dependent delta-6-desaturase having the substrate specificity of the delta-6-desaturase shown in SEQ ID NO:14, and
the above CoA-dependent delta-6-desaturase which has a preference for conversion of alpha linolenic acid compared to linoleic acid.
As this passage spans pp 6 and 7 of the application as filed, it has been referred to by the experts as the bridging paragraph. I will also use that description.
“CoA-dependent” describes a Δ6-desaturase that predominantly desaturates acyl-Coenzyme A (CoA) bound fatty acid substrates in the cytosolic acyl-CoA pool. SEQ ID NO: 13 is the polynucleotide which encodes the Δ6-desaturase from O. lucimarinus. SEQ ID NO: 14 is the polypeptide sequence encoded by SEQ ID NO: 13, that is, the CoA-dependent Δ6-desaturase polypeptide from O. lucimarinus. Of course, similar polynucleotides may either code for the same polypeptide or for a polypeptide with a similar sequence that preserves the identified biological function of interest. But of course, they may not.
As I have said, a substrate is the starting molecule upon which an enzyme acts. An enzyme may bind to and convert multiple substrates. Substrate specificity means the specific substrates, or the number of molecules, that an enzyme binds to at the catalytic site. There is a dispute as to the meaning of “the substrate specificity of the Δ6-desaturase shown in SEQ ID NO: 14”.
The experts have agreed that the words “the above” mean that the statement in the second bullet point of the bridging paragraph must be read together with the statement in the first bullet point of the bridging paragraph. Further, the feature in the first bullet point does not need to include the ALA preference feature disclosed in the second bullet point.
The conversion preference referred to means that the CoA-dependent Δ6 desaturase has greater desaturase activity on ALA relative to LA, that is it preferentially converts ALA to SDA over converting LA to GLA.
The experts have agreed that the bridging paragraph is the only place in which a conversion preference for ALA or, indeed, any conversion preference is disclosed in the body of the specification of the application as filed. Moreover, they have agreed that it is only disclosed in the limited context of, and as a feature of, a CoA-dependent Δ6-desaturase having the substrate specificity of the Δ6-desaturase shown in SEQ ID NO: 14.
The invention described in the bridging paragraph is specifically claimed in claims 1 and 2 respectively of the application as filed.
Page 7, lines 4 to 18 of the application as filed then describe a number of matters “[a]ccording to the invention…”. This provides:
According to the invention, the term “polynucleotide” relates to polynucleotides which comprise nucleic acid sequences which code for polypeptides with desaturase or elongase activity. The desaturase or elongase activities are preferably required for the biosynthesis of lipids or fatty acids. Especially preferably, they take the form of the following desaturase or elongase activities: Δ4-desaturase, Δ5-desaturase, Δ5‑elongase, Δ6-desaturase, Δ6-elongase or Δ12-desaturase. The desaturases and/or elongases are preferably involved in the synthesis of polyunsaturated fatty acids (PUFAs) and especially preferably in the synthesis of long-chain PUFAs (LCPUFAs). Suitable detection systems for these desaturase or elongase activities are described in the examples or in WO2005/083053. Especially preferably, the above-mentioned activities are, as regards substrate specificities and conversion rates, those of the respective enzymes from Ostreococcus lucimarinus. The specific polynucleotides according to the invention, i.e. the polynucleotides with a nucleic acid sequence as shown in SEQ ID NO: 1, 3, 5, 7, 9, 11, 13 or 15, were obtained from Ostreococcus lucimarinus.
Various desaturases and elongases, including Δ4-desaturase activity, Δ5-desaturase activity, Δ5-elongase activity, Δ6-desaturase activity, Δ6-elongase activity or Δ12-desaturase activity, are described. As indicated, it concludes:
Especially preferably, the above-mentioned activities are, as regards substrate specificities and conversion rates, those of the respective enzymes from Ostreococcus lucimarinus.
But in dealing with Δ6 desaturases, relevantly this is to be read in the context of the bridging paragraph.
The specification then states on p 7 line 20 and p 8 lines 15 to 20:
Therefore, polynucleotides according to the invention are in particular:
…
Polynucleotides which code for a polypeptide with Δ6-desasturase activity and which (i) comprise a nucleic acid sequence as shown in SEQ ID NO:13, (ii) comprise a nucleic acid sequence which codes for a polypeptide as shown in SEQ ID NO:14, (iii) comprise a nucleic acid sequence with at least 72% identity to one of the nucleic acid sequences of (i) or (ii), or (iv) a nucleic acid sequence of a fragment of a nucleic acid from (i), (ii) or (iii).
For convenience, and as the experts have done, I will refer to this as the page 8 paragraph.
The page 8 paragraph broadly describes polynucleotides which encode a polypeptide having Δ6-desaturase activity. According to the sequence listing, SEQ ID NO: 13 is a polynucleotide which encodes the Δ6-desaturase from O. lucimarinus. From a visual comparison, it would seem that SEQ ID NO: 14 is the polypeptide sequence encoded by SEQ ID NO: 13.
Sub-paragraph (i) of the page 8 paragraph provides for polynucleotides that incorporate SEQ ID NO: 13 and which code for a polypeptide having Δ6-desaturase activity.
Sub-paragraph (ii) provides for polynucleotides which code for a polypeptide having the amino acid sequence of SEQ ID NO: 14, which is a sequence that has Δ6-desaturase activity. But there would be a pool of polynucleotide sequences that could encode for a protein that has the activity of the protein of SEQ ID NO: 14, given the redundancy in the genetic code.
Sub-paragraph (iii) provides for polynucleotides which code for a polypeptide having Δ6-desaturase activity and which have at least 72% identity to those sequences of sub-paragraphs (i) or (ii). Percentage identity would be determined by performing a polynucleotide sequence alignment. On its face this provides for a broader range of polynucleotides than in (i) and (ii).
Finally, sub-paragraph (iv) provides for polynucleotides which code for a polypeptide having Δ6‑desaturase activity, but which comprise only a fragment of the sequences of (i), (ii) or (iii). This is also on its face a broader range of polynucleotides than in (i) and (ii).
In summary, sub-paragraphs (iii) and (iv) of the page 8 paragraph describe a broader range of polynucleotides than sub-paragraphs (i) and (ii) and the polypeptide that is coded for may have a range of differing properties. Now although the experts agree that, taken in isolation, the polynucleotide described at (iii) may not have the substrate specificity of the CoA-dependent Δ6‑desaturase shown in SEQ ID NO: 14 or the relevant conversion preference as described in the bridging paragraph, in my view the descriptions at page 7, line 13 and in the page 8 paragraph should be read in the context of the invention disclosed in the bridging paragraph. That is, read as a whole, the polynucleotides described in the page 8 paragraph are examples of polynucleotides coding for the invention described in the bridging paragraph.

Further, the invention is also said to relate to a process for the production of a substance which has the formula shown on pages 38 to 39. The formula covers an enormous array of chemical compounds and LC-PUFAs. This aspect of the invention is claimed in dependent claim 9 of the application as filed.
Figures 1 to 5 of the specification compare, respectively, the sequence alignment of Δ5- and Δ6-elongase, Δ4-desaturase, Δ5-desaturase, Δ6-desaturase and Δ12-desaturase amino acid sequences from O. lucimarinus and other unicellular algae (p 64 lines 5 to 20). Figure 4 relates to Δ6-desaturase.
Figures 6 to 10 show gas-chromatographic determination of fatty acids from yeasts which have been transformed with various plasmids (p 64 line 21 to p 65 line 5).
The specification concludes with five examples of “the present invention” (p 65 lines 7 and 8). Examples 1 to 3 relate to general methods of cloning, sequence analysis and lipid extraction. Example 4 relates to cloning and characterisation of elongase genes from O. lucimarinus. Example 5 relates to cloning and characterisation of desaturase genes from O. lucimarinus. Δ6‑desaturase with SEQ ID No: 14 (this did refer to SEQ ID No: 30 in the application as filed but “30” has now been corrected to “14”) is characterised and its sequence similarity is compared to Δ6 desaturases from other algae (p 72 line 15 to p 74 line 26). But the activity and substrate specificity is only provided for two Δ5-desaturases and a Δ12-desaturase and not the Δ6 desaturase (pp 74 to 77).
The application as filed ends in 15 claims. Claim 1 is the only independent claim. Claims 2 to 8 are product claims that depend on claim 1. Claims 9 to 11 are process claims that ultimately depend on claim 1. Claims 12 to 14 are use claims that ultimately depend on claim 1. Claim 15 is a product by process claim, dependent on claims 9 to 11.
Claim 1 of the application as filed is for:
A CoA-dependent delta-6 desaturase having the substrate specificity of the delta‑6 desaturase shown in SEQ ID NO: 14.
For convenience, and as the CSIRO has submitted, the “substrate specificity of the delta-6 desaturase shown in SEQ ID NO: 14” can be referred to as feature A. It is stipulated in the first bullet point of the bridging paragraph.
Dependent claim 2 of the application as filed is for:
The CoA-dependent delta-6 desaturase according to claim 1, wherein the desaturase has a preference for conversion of alpha linolenic acid compared to linoleic acid.
Again for convenience, a “preference for conversion of alpha linolenic acid compared to linoleic acid” can be referred to as feature B. It is stipulated in the second bullet point of the bridging paragraph. Accordingly, dependent claim 2 of the application as filed is for a CoA-dependent Δ6-desaturase having both feature A and feature B.
Dependent claim 3 of the application as filed is for:
An isolated polynucleotide comprising a nucleic acid sequence coding for the CoA-dependent Delta-6 desaturase according to claim 1.
Dependent claim 5 of the application as filed is relevantly for:
A host cell comprising the polynucleotide according to claim 3…
Dependent claim 9 of the application as filed is for:
A process for the production of a substance which has the structure shown in the general formula I hereinbelow:
where the variables and substituents are as follows:
R¹ =     hydroxyl, coenzyme A (thioester), lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylglycerol, lysodiphosphatidylglycerol, lysophosphatidylserine, lysophosphatidylinositol, sphingo base or a radical of the formula II
R² =     hydrogen, lysophosphatidylcholine, lysophosphatidylethanolamine, lysophosphatidylglycerol, lysodiphosphatidylglycerol, lysophosphatidylserine, lysophosphatidylinositol or saturated or unsaturated C₂-C₂₄-alkylcarbonyl,
R³ =     hydrogen, saturated or unsaturated C₂-C₂₄-alkylcarbonyl, or R² and R³ independently of one another are a radical of the formula Ia:
n = 2, 3, 4, 5, 6, 7 or 9, m = 2, 3, 4, 5 or 6 and p = 0 or 3;
and
wherein the process comprises the cultivation of (i) a host cell according to claim 5 or (ii) of a transgenic, nonhuman organism according to claim 7 or claim 8, under conditions which permit the biosynthesis of the substance.
A transgenic, nonhuman organism according to claim 7 or claim 8 is one which, inter-alia, comprises an isolated polynucleotide according to claim 3.
Dependent claim 12 of the application as filed is for the use of the polynucleotide, or a vector, host cell, transgenic, nonhuman organism comprising the polynucleotide, according to claim 3, for the production of an oil, lipid or fatty acid composition.
Finally, it is to be noted that no claim of the application as filed is dependent on claim 2. Further, the application as filed does not claim a process or use that relates to a CoA-dependent Δ6-desaturase having both features A and B. Claim 2 claims a product, being a CoA-dependent Δ6-desaturase having both features.

(b) The application as accepted

As I said earlier, the application as filed was amended prior to acceptance on three occasions. Let me note some aspects of the application as accepted incorporating such amendments.
The background to the invention described in the application as accepted is identical to that described in the application as filed. But importantly, the bridging paragraph at page 6 line 31 to page 7 line 2 of the application as filed, which defined the CoA-dependent ∆6-desaturase of the invention as having the substrate specificity of the ∆6‑desaturase shown in SEQ ID NO: 14 and a preference for conversion of ALA compared to LA, is not present in the application as accepted.
On page 6 line 31 to page 7b line 22 of the application as accepted, the bridging paragraph has been replaced with a description that defines the invention as being directed to a process for the production of substances or compounds of general formula I as follows:
A process for the production of a substance which has the structure shown in the general formula I hereinbelow:
where the variables and substituents for R¹, R² and R³ are as follows:
…
and
wherein the process comprises the cultivation of:
(i)        a host cell comprising an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO:14, wherein the polynucleotide is operatively linked to an expression control sequence; or
(ii)      a transgenic, nonhuman organism comprising:
a)        an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO:14,
b)        a vector comprising an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO:14, wherein the polynucleotide is operatively linked to an expression control sequence, or
c)        a host cell comprising an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO:14. wherein the polynucleotide is operatively linked to an expression control sequence,
under conditions which permit the biosynthesis of the substance, and
use of:
a)        an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO:14,
b)        a vector comprising an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO:14, wherein the polynucleotide is operatively linked to an expression control sequence,
c)        a host cell comprising an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO:14, wherein the polynucleotide is operatively linked to an expression control sequence, or
d)        a transgenic, nonhuman organism comprising:
(i)        an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO:14,
(ii)       a vector comprising an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO:14, wherein the polynucleotide is operatively linked to an expression control sequence, or
(iii)      a host cell comprising an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14. wherein the polynucleotide is operatively linked to an expression control sequence,
for the production of an oil, lipid or fatty acid composition.
The application as accepted defines the invention as being directed to a process for the production of substances or compounds of general formula I, wherein the process requires cultivation of a host cell or a transgenic non-human organism comprising a “polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent delta-6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14”. So, the invention is described as a process for production of a substance by reference to a general formula covering a large number of chemical compounds including LC-PUFAs comprising, and a use of, an isolated polynucleotide having a nucleic acid sequence coding for a CoA-dependent ∆6-desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14.
For convenience, the feature “at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14” can be referred to as feature C.
Now on its face, in my view a CoA-dependent ∆6-desaturase having feature C is a broader range of polypeptides than a CoA-dependent ∆6-desaturase having feature A, that is, the substrate specificity of the ∆6-desaturase shown in SEQ ID NO: 14. It describes CoA-dependent ∆6-desaturases that may have a range of different properties and may encompass a large number of variants which do not have feature A.
As CSIRO correctly points out, in light of the amendments to the bridging paragraph and the deletion of the corresponding claims, the application as accepted does not refer to feature A, feature B or feature A combined with feature B.
Further, the page 8 paragraph remains in the application as accepted and has not been modified from the application as filed.
Now as with the application as filed, the application as accepted describes that it relates to polynucleotides from O. lucimarinus which code for desaturases and elongases, and which can be employed for the recombinant production of polyunsaturated fatty acids. Further, the application as accepted describes that the object of the invention is to produce LC-PUFA in eukaryotic systems simply and inexpensively. But unlike the application as filed, the application as accepted claims to have achieved this object by provision of a process for the production of substances or compounds of general formula I. General formula I covers a vast array of chemical compounds and LC-PUFAs, including SDA, EPA, DPA, and DHA, either as free fatty acids or esterified onto a glycerol backbone.
The application as accepted has two independent claims that each refer to an isolated polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent ∆6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14.
Independent claim 1 of the application as accepted claims the process for production of compounds of general formula I, comprising the cultivation of a host cell or a transgenic non-human organism comprising a “polynucleotide comprising a nucleic acid sequence coding for a CoA-dependent ∆6-desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14” as described in the p 6 to p 7b extract set out earlier.
Independent claim 5 of the application as accepted claims the use of the isolated polynucleotide, a vector or host cell comprising the isolated polynucleotide, or a transgenic non-human organism comprising the isolated polynucleotide, vector, or host cell for the production of an oil, lipid or fatty acid composition as described in the p 6 to p 7b extract set out earlier.
The new description in the specification of the application as accepted (as described in the p 6 to p 7b extract set out earlier) corresponds to accepted process claim 1 and use claim 5.
Now claims 1 and 5 and the new description of the application as accepted define the CoA-dependent ∆6-desaturase the subject of the invention as having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14 (the O. lucimarinus ∆6-desaturase).
But this definition of the claimed CoA-dependent ∆6-desaturase does not equate with, and is not the same as, the requirement of the application as filed for a CoA-dependent ∆6-desaturase having the substrate specificity of the ∆6-desaturase shown in SEQ ID NO: 14.
The reason for this is that altering the nucleotide sequence may change the resulting polypeptide sequence, and could impact on the function of the resulting enzyme, therefore altering the substrate specificity of the enzyme. When nucleotide sequences are altered at multiple positions, the likelihood of the alteration impacting on the enzyme’s function increases. This is because the alteration may impact on the enzyme’s ability to recognise, bind and catalytically act upon particular substrates, therefore altering the enzyme’s substrate specificity.
Now I agree with CSIRO that although the application as filed provided a generic description of variants to the polynucleotide sequence encoding SEQ ID NO: 14, this was in the context of the sequence having one or more base substitutions, deletions and or insertions (page 8 line 35 to page 9 line 5); no specific polynucleotide variants were described. Also, it was stated that it was intended that the variants still encode for a polypeptide with the abovementioned biological activity of the respective starting sequence (page 9 lines 3 to 5). So, the invention could include variants, namely, polynucleotide sequences with at least 75% identity, provided that the variants had the same substrate specificity as the starting sequence.
And in this regard, the application as filed did not describe which nucleotides could be changed whilst still ensuring that the resulting nucleotide sequence having at least 75% identity to SEQ ID NO: 14 also coded for a polypeptide sequence that had the same substrate specificity of SEQ ID NO: 14.
Dr Singh gave evidence that he considered that a CoA-dependent ∆6-desaturase having at least 75% identity to a nucleotide sequence which coded for a polypeptide as shown in SEQ ID NO: 14 was broader than a CoA-dependent ∆6-desaturase having the same substrate specificity to SEQ ID NO: 14 because it included CoA-dependent ∆6-desaturases that did not have the same substrate specificity as SEQ ID NO: 14. I accept that evidence.
Indeed, to better illustrate the point that a CoA-dependent ∆6-desaturase having at least 75% identity to a nucleotide sequence which coded for a polypeptide as shown in SEQ ID NO: 14 does not equate with a CoA-dependent ∆6-desaturase having the substrate specificity of the ∆6‑desaturase shown in SEQ ID NO: 14, Dr Singh set out in a Table, which I have reproduced below, a comparison between the known fatty acid substrate specificities of three CoA-dependent ∆6-desaturases – Ostreococcus tauri (O. tauri), O. RCC809 and Micromonas pusilla (M. pusilla) CCMP1545 – and the O. lucimarinus CoA-dependent ∆6-desaturase shown in SEQ ID NO: 14.

Substrate specificity of CoA-dependent ∆6-desaturase variants of the CoA-dependent ∆6-desaturase shown in SEQ ID NO: 14

Before discussing the Table, let me note two lesser points. First, each percentage identity was calculated over an aligned region of a specified number of nucleotides. To so calculate, Dr Singh used the NCBI’s BLAST (Basic Local Alignment Search Tool) alignment program to align the nucleotide sequence of each of the O. tauri, O. RCC809 and M. pusilla ∆6-desaturases with the protein coding region of the nucleotide sequence encoding the polypeptide shown in SEQ ID NO: 14, applying the default settings of the standard package. Second, there is one error in the Table. In column 2, the reference to ω9-18:2^Δ9,12 should read ∆6, ∆9, omitting ∆12.
The results included in the Table were taken from three publications. The first publication was Yilmaz et al., “Determination of Substrate Preferences for Desaturases and Elongases for Production of Docosahexaenoic Acid from Oleic Acid in Engineered Canola”, Lipids (2017) 52 pp 207-222 (Yilmaz). The second publication was International patent application no. WO2010/057246 titled “Enzymes and Method for producing Omega-3 Fatty Acids” (246 Patent). And the third publication was Vaezi et al., “Identification and Functional Characterization of Genes Encoding Omega-3 Polyunsaturated Fatty Acid Biosynthetic Activities from Unicellular Microalgae”, Marine Drugs (2013) 11 pp 5116-5129 (Vaezi).
Column 1 of the Table identifies the fatty acid substrates the enzymes act on or bind to, to convert that fatty acid substrate to the fatty acid identified in column 2. This conversion was undertaken experimentally in yeast cells transformed with genetic constructs expressing the desaturase enzymes and which were fed the various fatty acid substrates. The conversion for each fatty acid substrate for each of the three ∆6-desaturase enzymes, compared to the O. lucimarinus ∆6-desaturase, that is, the polypeptide shown in SEQ ID NO: 14, is shown as a percentage of total fatty acids in columns 3 to 6.
Yilmaz demonstrated that the O. tauri ∆6-desaturase, which Dr Singh calculated has 79% identity over an aligned region of 1262 nucleotides to nucleotide sequence SEQ ID NO: 13 encoding the polypeptide sequence shown in SEQ ID NO: 14, has a substrate specificity for ETA, as it converts ETA to EPA (a ∆5-desaturase activity) (see Column 3 of the Table above and Table 2 of Yilmaz). In contrast, the O. lucimarinus ∆6-desaturase, that is, the polypeptide shown in SEQ ID NO: 14, had no substrate specificity for ETA, and did not convert ETA to EPA, as EPA was recorded in column 6 as “not detected” (see Table 7 of the 246 Patent). This result according to Dr Singh confirmed that whilst the O. tauri ∆6‑desaturase has “at least 75% identity” to the O. lucimarinus ∆6-desaturase of SEQ ID NO: 14, these two enzymes have a different substrate specificity. The nucleotide sequence for the O. tauri ∆6-desaturase gene and its characterisation were published before the priority date of the application as filed by Domergue et al “In vivo characterization of the first acyl-CoA ∆6 desaturase from a member of the plant kingdom, the microalga Ostreococcus tauri” Biochem J (2005) 389:483‑449 (Domergue), and its associated Gen Bank sequence listing (Gen Bank Accession No. AY646357.1).
Vaezi demonstrated that the O. RCC809 ∆6-desaturase, which Dr Singh calculated has 82% identity over an aligned region of 387-1833 nucleotides to nucleotide sequence SEQ ID NO: 13 encoding the polypeptide sequence shown in SEQ ID NO: 14, does not convert LA to GLA. In contrast, the O. lucimarinus ∆6-desaturase acts on LA.

I do not accept BASF’s position or Dr Stalker’s analysis on this aspect.
Dr Singh did not read the substrate specificity feature as only referring to the substrates that undergo Δ6-desaturation. This is because the meaning of “substrate specificity” is not limited to the primary catalytic activity of the enzyme. In his view, the term “substrate specificity” refers to the ability of an enzyme to recognise and bind to particular substrates, and to not bind to or act upon other related molecules within the context relevant to catalytic activity. That is, the phrase “the substrate specificity” is not limited to the substrates the enzyme binds to as part of its primary function. Rather, “substrate specificity” is understood in the art to refer to all substrates a particular enzyme can bind to and to potentially catalyse some type of reaction. He defined an enzyme’s specificity as the ability to discriminate between a substrate and a competing molecule. Moreover, he said that specificity was not limited to the enzyme’s primary catalytic activity. I accept that evidence providing that it is understood in the context of the biosynthesis contemplated in the application as filed, that is, relevant potential substrates.
Dr Singh said in the joint expert report that an enzyme can be categorised as a Δ6 desaturase by having a substrate specificity for at least one of the ω3 or ω6 fatty acids, namely, the ALA and/or LA substrates via insertion of a Δ6 double bond to produce the SDA and/or GLA products respectively. But the same enzyme may also act on shorter or longer fatty acid substrates in addition to ALA or LA, or other fatty acids having a C=C double bond at different positions of the respective fatty acid molecule. Further, a Δ6 desaturase may also introduce a C=C double bond at a different position, in addition to the Δ6 position. For this reason and as I have said, an enzyme described as a Δ6 desaturase may have substrate specificity for other different fatty acid substrates. Two Δ6 desaturases that act only on ALA and LAs substrates would have the same (identical) substrate specificity. But as I have said, assume that the second enzyme of the two Δ6 desaturases can also act on the substrate ETA to introduce a C=C double bond at the 5th carbon atom from the carboxyl end (a Δ5 desaturase activity). It would have a different substrate specificity. The second enzyme would have a substrate specificity for three substrates (ALA, LA and also ETA), whereas the first enzyme would only have a substrate specificity for two substrates (ALA and LA). The Δ6 desaturase from O. tauri has activity on ALA, LA as well as ETA while the Δ6 desaturase from O. lucimarinus (SEQ ID NO: 14) does not have activity on ETA. Hence these two enzymes do not have the same substrate specificities. Further, some enzymes can catalyse more than one type of reaction. This is exemplified by O. tauri and Micromonas Δ6 desaturases. Further, O. tauri Δ6 desaturase can also insert a Δ6 double bond in the monounsaturated fatty acid oleic acid to produce isolinolenic acid. Further, unlike O. tauri and Micromonas Δ6 desaturases, SEQ ID NO: 14 has no detectable specificity for the substrates ETA and ETrA.
Now the bridging paragraph refers to the enzyme as being a CoA-dependent Δ6-desaturase because that is the enzyme’s primary catalytic activity. But the CoA-dependent Δ6- desaturase may be able to undertake other catalytic activities, for example a Δ5-desaturation at the Δ5 position. If so, it would still be referred to as a Δ6-desaturase because that activity is the enzyme’s primary function. However, according to Dr Singh, the reference to “the substrate specificity” in the bridging paragraph was not limited to the substrates the enzyme binds to as part of its primary catalytic activity. I must say that I agree with Dr Singh.
Further, Dr Singh also considered that Dr Stalker’s reading of the bridging paragraph gave the words “having the substrate specificity of the Δ6 desaturase shown in SEQ ID NO: 14” no work to do. Dr Stalker read the feature as referring to any CoA-dependent Δ6-desaturase rather than “a CoA-dependent Δ6 desaturase having the substrate specificity of the Δ6 desaturase shown in SEQ ID NO: 14”. Accordingly, Dr Stalker’s reading gives no work to do to the qualifier.
Further, Dr Singh disagreed with Dr Stalker’s interpretation that the phrase “the substrate specificity” does not mean the same relevant substrate specificity. This is because the substrate specificity feature specifically refers to the CoA-dependent Δ6-desaturase “having the substrate specificity of the Δ6 desaturase shown in SEQ ID NO: 14” (my emphasis). Dr Singh understood the word “the” to denote a specific or particular effect. For this reason, he read the substrate specificity feature as saying that the invention is directed to a CoA-dependent Δ6-desaturase having “the same” or “the specific” or “the identical” substrate specificity of “the Δ6 desaturase shown in SEQ ID No:14”. And this therefore requires a comparison with the specific substrates that the Δ6-desaturase shown in SEQ ID NO: 14 binds to, being the substrates that the enzyme binds to as part of its primary activity (being a Δ6 desaturase) and any substrates that enzyme binds to as part of any side reactions, related reactions or bifunctional reactions, but of course relevant to the biosynthetic pathway in question.
Further, Dr Singh said that although a Δ6-desaturase will predominantly desaturate LA or ALA to produce GLA or SDA respectively, it has also been shown to bind to oleic acid (OA) to produce isolinoleic acid through desaturation at the sixth carbon. For example, a recent paper detailed a study showing that the Δ6-desaturase from Phytophthora citrophthora transformed in Perilla seeds bound to OA, LA and ALA to produce isolinoleic acid, GLA and SDA respectively (Lee et al, High accumulation of γ-linolenic acid and Stearidonic acid in transgenic Perilla (Perilla frutescens var. frutescens) seeds, BMC Plant Biology (2019) 19:120). Further, the O. tauri Δ6-desaturase has also been reported to have Δ6-desaturase activity on OA to produce isolinoleic acid; see Domergue that I cited earlier. Accordingly, Dr Singh did not read the substrate specificity feature as encompassing only the substrates LA or ALA.
In my view, the expression “the substrate specificity of the ∆6-desaturase shown in SEQ ID NO: 14” (my emphasis) serves to describe the specificity which is unique or specific to that particular polypeptide relevant of course to the biosynthetic context one is considering.
The requirement that the CoA-dependent ∆6-desaturase have the substrate specificity of the ∆6-desaturase shown in SEQ ID NO: 14 defines and limits the substrate specificity of the CoA ∆6-desaturase described and claimed to those which possess the particular specificity of the ∆6-desaturase shown in SEQ ID NO: 14.
Now a ∆6-desaturase will at least have a substrate specificity for ALA and/or LA. This is its primary catalytic activity. But a ∆6-desaturase may also have substrate specificity for different fatty acid substrates.
Dr Stalker wrongly equated the primary catalytic activity of any ∆6-desaturase (i.e. acting on ALA and/or LA) with the substrate specificity of the ∆6-desaturase shown in SEQ ID NO: 14. Primary catalytic activity is not the same as “the substrate specificity”.
Further, Dr Stalker’s reading would render the words “having the substrate specificity of the ∆6-desaturase shown in SEQ ID NO: 14” redundant. This is because a “CoA-dependent Δ6 desaturase”, without more, already describes any “enzyme that functions as a ∆6-desaturase and thus has an affinity for ALA and/or LA.”
Further, Dr Stalker’s interpretation ignores the qualifying nature of the word “the”.
Further, although a ∆6-desaturase will predominantly desaturate LA and/or ALA, it may also have ∆6-desaturase activity on OA to produce isolinoleic acid by desaturation at the sixth carbon. Accordingly, even if this expression referred only to the ∆6-desaturase activity of an enzyme, it would not be limited to the substrates LA and ALA.
In the context of the application as filed, in my view “the substrate specificity” refers to all of the fatty acid substrates within the LC-PUFA pathway that the enzyme binds to.
As to the CoA-dependent Δ6-desaturase having the relevant substrate specificity, it must have all of the fatty acid substrates of the Δ6-desaturase shown in SEQ ID No: 14 within the LC-PUFA pathway. Examples of such substrates for Δ6-desaturases within the LC-PUFA pathway include LA, ALA, OA. But it is not required to have those substrates of the Δ6-desaturase shown in SEQ ID No: 14 that do not relevantly form part of the LC-PUFA pathway (if any); such substrates would be understood by the skilled addressee to be extraneous to the relevant context.
Now I agree with CSIRO that it is not for it to advance a practical explanation as to why the patent applicant chose to limit its description of the invention in this manner. The specification of a patent is a document in words of the patentee’s own choosing, usually drafted with highly skilled and expensive advice. A patentee may have good reason for introducing a limitation into a claim, including to avoid arguments in relation to prior art. For example, I note that Domergue discloses a Δ6-desaturase that binds to and converts four substrates in the LC-PUFA pathway, including ALA and LA. It may be speculated that if the first bullet point of the bridging paragraph did no more than describe a Δ6-desaturase that binds to and converts ALA and LA, the invention may have been vulnerable to attack on the ground of lack of novelty.
BASF’s construction involves impermissibly ignoring the limitation the patentee chose to introduce into the claim, that is, “having the substrate specificity of the Δ6-desaturase shown in SEQ ID NO: 14”.
Further, BASF’s construction involves adding a gloss to the words of the disclosure such as core or primary substrate specificity.
Further, on BASF’s construction, a Δ6-desaturase acting on two substrates within the LC-PUFA pathway would have the same substrate specificity as a Δ6-desaturase acting on four substrates in that pathway because they have two common substrates. But there is no cogent reason that would support such a conclusion.
In summary, I reject BASF’s construction involving “substrate specificity”. This conclusion is also sufficient to dispose of BASF’s tertiary argument.

(b) Preference for conversion of ALA compared to LA

In addition to the CoA-dependent Δ6-desaturase having the same substrate specificity (in context) as the Δ6-desaturase shown in SEQ ID NO: 14, the application as filed also states that the CoA-dependent Δ6-desaturase is to have a preference for conversion of the ω3-fatty acid, ALA compared to the ω6-fatty acid, LA. That is, it operates preferentially in the ω3 pathway, rather than the ω6 pathway shown in the schematic that I set out earlier in my reasons.
Now no definition of “preference” is provided in the application as filed. The only two references to “preference” in the application as filed are on page 7 line 1 and claim 2.
But it is not in dispute that the term “preference” in this context refers to a quantitative comparison of the rate of conversion of ALA to SDA relative to the conversion of LA to GLA. Preference refers to greater desaturase activity on ALA relative to LA. On this basis, in any relevant study involving the claimed CoA-dependent Δ6-desaturase, it would be expected that the resulting product would contain a higher amount of SDA than GLA, relative to the amount of ALA and LA added in the study, as a result of the claimed CoA-dependent Δ6-desaturase’s preference to convert ALA into SDA.
Now I should say at this point that the concepts of conversion preference and conversion rate should not be confused. The former deals with a comparison of rates. Let me return to the application as filed.
The application as filed describes a CoA-dependent Δ6-desaturase having the substrate specificity of the Δ6-desaturase shown in SEQ ID NO: 14 and also having a preference for conversion of ALA compared to LA. This is because of the use of the conjugative “and” at page 6 line 34 which refers to ALA conversion preference as being in addition to the requirement of identical substrate specificity. This conclusion is further supported by the reference to “the above” CoA-dependent Δ6-desaturase at page 7 line 1, which defines the feature of ALA conversion preference by reference to, and combined with, the previously mentioned feature of substrate specificity.
Now in the context of the bridging paragraph, the statement disclosed in the first bullet point can be independent of the statement disclosed in the second bullet point. So, the statement disclosed in the first bullet point, ie, the substrate specificity of the Δ6 desaturase shown in SEQ ID NO: 14 does not need to include the ALA preference feature disclosed in the second bullet point. But the words “the above” entail that the statement disclosed in the second bullet point needs to be read in context with the statement in the first bullet point.
The experts were asked to consider the application as filed, and explain whether they considered that it disclosed a CoA dependent Δ6-desaturase that preferentially converts ALA compared to LA. They gave the following evidence. Preference in context is a qualitative comparison of the rate of conversion of ALA to SDA relative to the rate of conversion of LA to GLA. On this basis, in any expression test involving the claimed CoA-dependent Δ6 desaturase, they would expect that the enzymatic reaction would produce a higher amount of SDA than GLA. Now in the application as filed, a Δ6 desaturase that has a preference for ALA as a substrate does not define the degree of preference for ALA as compared to LA as a substrate. And there is no specific data in the application as filed showing the degree of preference for the Δ6 desaturase shown in SEQ ID NO: 14 to convert ALA over LA. Further, there is no data in the application as filed showing preference for ALA as a substrate for any of the other O. lucimarinus sequences disclosed. The only mention of the words “preference for ALA” in the application as filed occurs in the bridging paragraph and claim 2. There is a reference to “[e]specially preferably” on page 7 line 13, and in that context the page 8 paragraph refers to Δ6 desaturase activity. But those descriptions do not broaden the scope of the disclosure. The page 8 paragraph is to be read in the context of the subject of the invention of the application as filed i.e. a CoA-dependent Δ6 desaturase having the substrate specificity of SEQ ID NO: 14 (claim 1) and that this CoA-dependent Δ6 desaturase may also have a substrate preference for ALA over LA (claim 2).
Further, claim 1 of the application as filed claims “a CoA-dependent Δ6 desaturase having the substrate specificity of the Δ6 desaturase shown in SEQ ID NO: 14”. That is claim 1 claims only the feature of substrate specificity.
Claim 2 of the application as filed claims “the CoA-dependent Δ6 desaturase according to claim 1, wherein the desaturase has a preference for conversion of [ALA] compared to [LA]”. That is, claim 2 claims both of the features of substrate specificity and ALA conversion preference.
Accordingly, the claims of the application as filed are consistent with the interpretation that the preference for conversion of ALA compared to LA is a feature of a CoA-dependent Δ6-desaturase that also has the substrate specificity of the Δ6-desaturase shown in SEQ ID NO: 14.
The experts were asked to consider whether the application as filed disclosed a CoA-dependent Δ6-desaturase that does not have the substrate specificity of the Δ6-desaturase shown in SEQ ID NO: 14 but which preferentially converts ALA compared to LA. They agreed that there was no such express disclosure.
In summary, the application as filed discloses a CoA-dependent Δ6-desaturase that has the ALA conversion preference which also has the same substrate specificity as the CoA-dependent Δ6-desaturase shown in SEQ ID NO: 14. There is no disclosure in the application as filed of a CoA-dependent Δ6-desaturase that has the ALA conversion preference without such CoA-dependent Δ6-desaturase also having the same substrate specificity of the CoA-dependent Δ6-desaturase shown in SEQ ID NO: 14.
The experts were also asked to consider whether a CoA-dependent Δ6-desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14 and which preferentially converts ALA compared to LA was disclosed in the application as filed.
Dr Singh’s opinion was no. The application as filed is limited to a CoA dependent Δ6 desaturase both having the substrate specificity shown in SEQ ID NO: 14 and having a preference for ALA over LA as a substrate. There are 2 scenarios that he said were worth considering here.
First, it is possible that a Δ6 desaturase having at least 75% homology at the DNA level to the Δ6 desaturase shown in SEQ ID NO: 14 also has the substrate specificity of the Δ6 desaturase shown in SEQ ID NO: 14.
Second, it is also possible that a Δ6 desaturase, where the DNA coding region has at least 75% homology at the DNA level to the sequence that codes for the polypeptide shown in SEQ ID NO: 14, exhibits a more relaxed substrate specificity than the Δ6 desaturase shown in SEQ ID NO: 14. Examples of Δ6 desaturases cited in Dr Singh’s Table that I have reproduced earlier illustrate this scenario. For example, the O. tauri Δ6 desaturase has 79% nucleotide homology to SEQ ID NO: 13 (coding for the polypeptide shown in SEQ ID NO: 14) of the application as filed. The O. tauri Δ6 desaturase can also convert the substrate ETA to EPA, whereas the polypeptide coded for by SEQ ID NO: 13 does not perform that function. It is noteworthy that conversion of ETA to EPA is in the context of the entire PUFA pathway shown in the biosynthetic pathway schematic that I have set out earlier. In fact, a single amino acid alteration could alter the function and thereby change the substrate specificity of an enzyme. So, it is also possible that two enzymes can have different substrate specificities and have a 75% homology to each other at the DNA level. Dr Singh said that due to the existence of this scenario, a 75% comparative homology at the DNA level for a Δ6 desaturase cannot be taken to mean that such a Δ6 desaturase has the substrate specificity of the Δ6 desaturase shown in SEQ ID NO: 14 in the application as filed. I must say that I had no reason to doubt such evidence.
One can contrast all of this with the proposed amendments.
The amendments, by seeking to insert new claims 2 and 7 into the application as accepted, would claim a CoA-dependent Δ6-desaturase:
(a) having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14 (see claims 1 and 5 (now claim 6) of the application as accepted); and
(b) that preferentially converts ALA compared to LA (proposed new claim 2, when read together with claim 1, of the application as accepted).
But the second matter is new matter that was not originally disclosed in the application as filed. This is because this feature of a CoA dependent Δ6-desaturase that preferentially converts ALA compared to LA was only ever disclosed in the context of a CoA-dependent Δ6-desaturase that has the substrate specificity of the Δ6-desaturase shown in SEQ ID NO: 14.
By contrast, a CoA dependent Δ6-desaturase having the substrate specificity of the Δ6-desaturase shown in SEQ ID NO: 14 and the ALA conversion preference as claimed in the application as filed is not the same as a CoA-dependent Δ6-desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14 and having the ALA conversion preference (as claimed in new claims 2 and 7).

So new claims 2 and 7 would be seeking to claim a CoA-dependent Δ6-desaturase that was not disclosed in the application as filed. This is because the application as filed was limited to a CoA-dependent Δ6-desaturase that has both the identical substrate specificity and ALA conversion preference features, whereas new claim 2 is seeking to claim a CoA-dependent Δ6-desaturase that has the ALA conversion preference only, together with having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14, which is not the same as identical substrate specificity.
Let me say something about the amendment which seeks to insert the following description into the body of the specification of the application as accepted (at page 7b):
According to an embodiment of the abovementioned process and use, the CoA‑dependent desaturase preferentially converts [ALA] compared to [LA].
This amendment is seeking to describe a CoA-dependent Δ6-desaturase having this preference for ALA, but where the CoA-dependent Δ6-desaturase has at least a 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14 (as opposed to having the substrate specificity of SEQ ID NO: 14).
But given that substrate specificity and at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14 are different, this amendment, by seeking to insert this new paragraph into page 7b of the application as accepted, would be seeking to add new material that was not disclosed in the application as filed in terms of amounting to an impermissible intermediate generalisation.

(c) Page 8 paragraph

Page 7, line 20 states, “Therefore, polynucleotides according to the invention are in particular:...”. Page 7 line 21 to page 8 to line 27 describe polynucleotides that are preferably expressed in the host cell or transgenic non-human organism encoding the suite of enzymes involved in the production of fatty acids, specifically DHA, as per the pathway shown in the figure for the schematic representation of the conversion of LA or ALA. Specific polynucleotides described include SEQ ID NO: 1, 3, 5, 6, 7, 9, 11, 13, 15 or 16 encoding a suite of enzymes, including a Δ4-desaturase, a Δ5-desaturase, a Δ5-elongase, a Δ6-desaturase a Δ6-elongase and a Δ12 desaturase. Although elongases and desaturases other than Δ6-desaturases are referred to in this paragraph and elsewhere in the specification, the invention as claimed in both the application as filed and the application as accepted is limited to certain Δ6-desaturases.
In relation to the Δ6-desaturase, the page 8 paragraph is “therefore” an example of “the invention”. In my view this entails that the polynucleotides in part (iii) of the page 8 paragraph must have the required substrate specificity coupled with the ALA conversion preference as described in the bridging paragraph.
The skilled addressee does not read the page 8 paragraph in isolation. The page 8 paragraph discloses the polynucleotides “according to the invention” (application as filed, page 7, line 20). The page 8 paragraph does not broaden the scope of the invention disclosed. It is to be read in the context of the bridging paragraph.
Now Dr Stalker gave evidence that parts (iii) and (iv) of the page 8 paragraph provide a broader range of polynucleotides than (i) and (ii), provided that the Δ6-desaturase activity is preserved. But polynucleotides within parts (iii) and (iv) must also preserve the required substrate specificity and ALA conversion preference features as disclosed in the bridging paragraph.
The descriptions which follow on pages 9 to 11 and page 30 lines 1 to 2 and 25 to 28 of the application as filed are subject to the same limitation.
Further, in relation to a Δ6-desaturase which has the ALA conversion preference, the application as filed only discloses and claims a Δ6-desaturase as having this feature, in the context of the Δ6-desaturase also having “the substrate specificity” feature of the Δ6- desaturase shown in SEQ ID NO: 14. That is, the application as filed does not disclose the ALA conversion preference independently of the required substrate specificity feature.
Now although Dr Stalker acknowledged the conjugative “and” in the bridging paragraph at the end of the first bullet point, insufficient attention was given to the reference to “the above” at page 7 line 1. But the reference to “the above” defines the ALA conversion preference feature by reference to, and combined with, the previously mentioned feature of the required substrate specificity.
Now BASF contends that read in isolation, there is a disclosure in the page 8 paragraph of a polynucleotide coding for a Δ6 desaturase having the amino acid sequence disclosed as SEQ ID NO: 14 and of certain polynucleotides within specified ranges of sequence homology. Further, it contends that the second bullet point of the bridging paragraph discloses that the SEQ ID NO: 14 polypeptide preferentially converts ALA compared to LA.
Therefore, the skilled addressee understands from the application as filed that:
(a)       Δ6 desaturases will act in LC-PUFA biosynthetic pathways; and
(b)       as a subset of the above, Δ6 desaturases will preferentially convert ALA to LA; and
(c)       polynucleotides having the requisite homology (75% identity) will likely produce polypeptides that preserve those biological functions.
But BASF impermissibly seeks to strip the conversion preference (feature B) from the context in which it is disclosed in the bridging paragraph, by attempting to describe it as a property of SEQ ID NO: 14 and therefore combine feature B with the required sequence homology (feature C). But feature C is not coterminous with the required substrate specificity (feature A).
Further, BASF’s argument proceeds on a flawed foundation.
The conversion preference was to be applied to the CoA-dependent Δ6 desaturase having the substrate specificity of the Δ6 desaturase shown in SEQ ID NO: 14. This feature was not directly referable to the Δ6 desaturase shown in SEQ ID NO: 14 itself.
The bridging paragraph does not disclose the conversion preference of SEQ ID NO: 14. The skilled addressee would not understand from the second bullet point of the bridging paragraph or any part of the application as filed that one of the properties of SEQ ID NO: 14 itself is that it will preferentially convert ALA compared to LA.
First, there is no data in the application as filed to support that proposition.
Second, the bridging paragraph discloses a class of enzymes that answer the description of “a CoA-dependent Δ6-desaturase having the substrate specificity of the Δ6-desaturase shown in SEQ ID NO: 14.” Accordingly, the disclosure of the class of enzymes described in the first bullet point is not a disclosure of the conversion preference for SEQ ID NO: 14. That is, the substrate specificity of the class of enzymes described in the first bullet point is not determinative of the conversion preference of those enzymes.
Third, such a class is narrowed in the second bullet point. But it does not disclose a property of SEQ ID NO: 14.
Fourth, the difference in language between the first bullet point and the second bullet point is significant. It does not describe the conversion preference by reference to the Δ6-desaturase shown in SEQ ID NO: 14.
In summary, in my view the page 8 paragraph does not disclose the invention the subject of the proposed amendment. The specific processes and uses BASF now seeks to disclose and claim with respect to a CoA dependent Δ6-desaturase with feature C combined with feature B is, at best, a “subset” of the page 8 paragraph considered in isolation. But in my view the skilled addressee would not have read the page 8 paragraph in isolation in the application as filed. And nor would they have read the second bullet point of the bridging paragraph in isolation. In my view, a person skilled in the art would not so read the page 8 paragraph out of context as Dr Stalker has done.
So, the page 8 paragraph of the application as filed does not disclose a CoA-dependent Δ6-desaturase:
(a)       having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14 (proposed new claim 6); and
(b)       that preferentially converts ALA compared to LA (proposed new claim 2, when read together with claim 1, of the application as accepted).
Accordingly, BASF’s primary argument should be rejected.

(d) Other matters

Is anything left of the secondary argument? I do not think so. In my view one cannot diminish the significance of the bridging paragraph as BASF has sought to do. Now I accept that the bridging paragraph discloses the conversion preference. But as I say, that was in a specific and in substance cumulative context.
In summary, BASF by the amendment seeks to take the conversion preference feature well outside the context of the application as filed. In the application as filed it was limited to a CoA dependent Δ6 desaturase that also had the substrate specificity of SEQ ID NO: 14. So, it was dealing with such a desaturase that had both features, not the conversion preference feature alone. Further, the preferments were to be understood in the context of the bridging paragraph. Further, the page 8 paragraph did not describe a broader invention. Rather, the polynucleotides were those which coded for the polypeptides of the invention described in the bridging paragraph.
So, when one considers the application as filed read as a whole through the eyes of the skilled addressee, it relevantly disclosed the invention as:
(a) a CoA-dependent Δ6 desaturase with the relevant substrate specificity; and
(b) a CoA-dependent Δ6 desaturase having both the relevant substrate specificity and the conversion preference.
Contrastingly, the proposed amendments take the conversion preference feature (feature B) and add it to the feature of a CoA-dependent Δ6 desaturase having at least 75% identity to a nucleotide sequence which codes for a polypeptide as shown in SEQ ID NO: 14 (feature C). But this was not disclosed in the application as filed. To put it bluntly, the relevant substrate specificity (feature A) and feature C are not the same.
Moreover, even if the relevant disclosure in the application as filed is not limited to the invention, the amendment is still not allowable. Even though the page 8 paragraph or other stray references may include or cover the matters in the application as proposed to be amended, it does not disclose those matters for the purpose of s 102(1). It does not disclose the specific processes and uses it now seeks to disclose and claim with respect to feature C combined with feature B.
If I have not already made it clear, you cannot just take the page 8 paragraph as free-standing and add to it feature B. To do so you are decontextualising both. Both are to be read in the context of the bridging paragraph. And nor is it permissible to just take the page 8 paragraph as free-standing and to say that the page 8 paragraph together with feature B is just a sub-set. Again that is impermissibly decontextualising.
Accordingly, the proposed amendments constitute an impermissible intermediate generalisation. Let me conclude by making the following other points to the extent that I have not already done so.
First, BASF has said that features A and B from the bridging paragraph are not combined in the sense of a combination of physical integers. They each describe properties of an individual polypeptide. Now this may be accepted. But in my view, so to accept does not deny the force of CSIRO’s point concerning intermediate generalisation.
Second, BASF also says that features A and B do not co-operate to achieve some unitary effect or outcome. But even if this be so, that does not deny the proposition that the re-addition of feature B amounts to an impermissible intermediate generalisation.
Third, BASF has also sought to place some emphasis on the passage in the application as filed which begins “Especially preferably…” (p 7 line 13). But in the context of Δ6 desaturases, this is not broadening from the bridging paragraph. And in terms it is not referring to the conversion preference in the second bullet point of the bridging paragraph.
Fourth, let me say something further concerning claims. BASF says that although claims of the complete specification as filed can form part of the disclosure, they do not limit the scope of the disclosure. I agree. But this does not really take BASF anywhere.
Fifth, BASF says that the amendments are narrowing rather than broadening. As I have said, seen in the context of the application as accepted, they are narrowing. But seen in the context of a comparison with the application as filed, that is not a satisfactory description. In any event, I prefer not to use a narrowing/broadening discriminant. That is not the language of s 102(1).
Finally, let me make one other observation. I have not engaged in any discussion as to whether the proposed amendments would have also been disallowed under the old version of s 102(1). I prefer to avoid any retrospective hypothetical.
The appeal is allowed. The proposed amendments are impermissible. I will make the necessary orders.

I certify that the preceding three hundred and forty eight (348) numbered paragraphs are a true copy of the Reasons for Judgment herein of the Honourable Justice Beach.

Associate:

Dated: 12 March 2020

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