DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Terminal Disclaimer Proper
The terminal disclaimer filed on 07/17/2025 disclaiming the terminal portion of any patent granted on this application which would extend beyond the expiration date of any patent granted on Application Number 17775168 has been reviewed and is accepted. The terminal disclaimer has been recorded.
Status of the Claims
The amendment filed 07/14/2025 has been entered. Claims 1, 4, and 6-29 are pending and are examined in this office action.
Rejection that are withdrawn
Objection to claim 20 is withdrawn in light of applicant’s amendment of claims now deleting the recitation of claim 1.
Obviousness type nonstatutory double patenting over copending Application No. 17775168 has been withdrawn in light of applicant’s terminal disclaimer filed on 07/17/2025.
Response to Argument on declaration under 37 CFR 1.132
The declaration under 37 CFR 1.132 filed on 07/18/2025 and 07/14/2025 is insufficient to overcome the rejection of claims 1, 4, and 6-29 based upon a specific reference applied under 35 U.S.C. 103 as set forth in the last Office action because: it fails to set forth facts.
Response to Declaration of Dr. Jerry Hjelle
Declaration of Dr. Jerry Hjelle argues a person skilled in the art at the time would not read Aziz as disclosing any relevant information, assay results, or other data regarding the toxic effects of Cry1Da against insects, let alone specifically lepidoptera. Applicant asserts the disclosure is a narrow comparison of two different protein expression systems---E. coli and Baculovirus. Dr. Jerry Hjelle asserts reasonable conclusion of the disclosure is that a fragment of the native Cry1Da gene can be expressed in either or both of these in vitro fermentation production systems (Dr. Hjelle Declaration, page 2, last paragraph). Dr. Jerry Hjelle argues stated differently, the disclosure of Aziz provides no meaningful data on either the level toxicity or spectrum of insecticidal activity, and, as a result, it would not be read by a person skilled in the art as a promising toxin for expression in transgenic plants in order to obtain useful insect pest control (Dr. Hjelle Declaration, page 3, first paragraph).
Dr. Jerry Hjelle argued Baum leads a person skilled in the art looking for a Cry1Da with improved toxicity and spectrum away from using either the full length or the truncated sequence of the wild-type Cry1Da of Aziz for the following reasons: (Dr. Hjelle Declaration, pages 3 and 4).
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Dr. Jerry Hjelle argues a person skilled in the art looking for a Cry1Da with improved spectrum or toxicity who read Baum would not find motivation to design a nucleic acid molecule for expressing the truncated version of the wildtype Cry1Da disclosed in Aziz. Dr. Jerry Hjelle argues rather a person skilled in the art would be motivated to perform amino acid substitutions on the full length wild-type Cry1Da protein to enhance spectrum of activity. Dr. Jerry Hjelle argues even if a person of skill in the art considered a truncation, there is no teaching that would motivate a person art to choose the specific 625 aa fragment of Aziz over other possible truncations (Dr. Hjelle Declaration, pages 5, paragraph 3).
Dr. Jerry Hjelle argues during transformation vector construction, Applicants modified the protein coding sequence of the native Cry1Da gene by truncating the native sequence to produce the 625 amino acid N-terminal portion of the protein containing Domains I, II and III of the toxin. Dr. Jerry Hjelle argues in stark contrast to what was expected by a person skilled in the art from the abovementioned prior art, as shown in Dr. Camila's Declaration of November 14, 2024, a truncated variant consisting of the 625 amino acid N-terminal portion of the protein of the native Cry1Da proteins had LC50 values (and growth inhibition at even lower doses) that could easily provide a commercially useful lepidoptera inhibitory spectrum and activity (Dr. Hjelle declaration, page 5, paragraphs 4-5).
Dr. Jerry Hjelle argues in stark contrast to what was expected by a person skilled in the art from the abovementioned prior art, as shown in Dr. Camila's Declaration of November 14, 2024, a truncated variant consisting of the 625 amino acid N-terminal portion of the protein of the native Cry1Da proteins had LC50 values (and growth inhibition at even lower doses) that could easily provide a commercially useful lepidoptera inhibitory spectrum and activity (Dr. Hjelle Declaration, pages 5-6, last and first paragraph).
The Declaration filed under 37 C.F.R. § 1.132 by Dr. Hjelle have been fully considered but they are not persuasive since although Dr. Hjelle asserts Aziz et al. express fragment of the native Cry1Da gene in E. coli and Baculovirus, Dr. Hjelle argues there is no meaningful data on either level toxicity or secretum of insecticidal activity. Generating such meaningful data will be within reach of a skilled in the art. Aziz et al. discloses the information is dated 03/2018 before the effective date of filling of the invention in https://www.researchgate.net/publication/323627747_Bacillus_thuringiensis_strain_aizawai_active_core_crystal_toxin_protein_1D_gene_partial_cds_ACCESSION_JQ910166, accessed 03/07/2025 that their data showed the active core cry1D gene sequence from genomic DNA of Bacillus thuringiensis subsp. Aizawai and the gene coded for the active form of Cry1D crystal protein and has been proven toxic to its specific targeted-insect pests which were Spodoptera litura and Spodoptera exigua (see enclosed snippet and the entered accession JQ910166.1 included with the website). The previously attached locus of the ResearchGate website JQ910166.1 encodes SEQ ID NO:3. For this reason it was known to have been effect to insects from genus Spodoptera.
Regarding Baum’s teaching of wildtype Cry1Da proteins does not exhibit commercially useful Lepidopteran inhibitory spectrum and improved Lepidopteran inhibitory activity, and Baum does not provide definition of commercially useful activity or spectrum as the toxicity (potency) of the specific expressed Bt toxin and the dose. Finding specific toxicity and dose of a known protein taught by Aziz et al. is routine procedure for a skilled in the art. Aziz et al. teaches the isolation source of their disclosed locus AFK29089 is from commercial Bt based insecticide (see description in “source of the locus AFK29089). Furthermore, there is no indication that the applicants have applied any inventive step that has changed the toxicity and dose of the disclosed protein by Aziz et al.
Furthermore, having a higher toxicity for the fragment compared to the full-length protein is not surprising or unexpected because this trend/characteristic was already known in the prior art Pardo-Lopez et al. (see analysis above), and the specific amount of activity for SEQ ID NO: 3 is an inherent property of SEQ ID NO: 3 which is specifically taught by Aziz et al. Furthermore, the activity of LC50 concentrations against Spodoptera frugiperda, D. sacharalis, and Helicoverpa zea at 0.13. 0.15, and 0.92 ug/cm2, respectively would have been inherent property of the toxin disclosed by Aziz et al.
Regarding argument on Baum teaching away from the actual commercial utility of the native Cry1Da protein in controlling H. zea, Aziz et al. teaches truncated protein of Baum and Baum teaches their disclosed insecticidal proteins for example SEQ ID NO: 2 can be truncated forms wherein one or more amino acids are deleted from the N-terminal end, C-terminal end, the middle of the protein, or combinations thereof without a loss of insect inhibitory activity and these fragments should retain the insect inhibitory activity of the parent engineered insecticidal protein (page 7, paragraph 0080). Baum teaches engineered toxic protein contain amino acids substitution addition or deletions as compared to Baum’s SEQ ID NO: 2 which have showed unexpectedly improved insecticidal activity and enhanced insecticidal spectrum against Lepidopteran insect pests including H. zea (page 1, paragraph 0008). The deletion is a form of truncation. Furthermore, applicant is basing the argument on single art when the obviousness is determined by combination of arts for example Pardo-Lopez et al. teaches deletion of small fragments from amino-terminal region led to improved toxicity or overcome resistance representing interesting alternatives for insect pest control (page 589, Abstract and page 593).
Furthermore, as previously discussed evidence in prior office action that use of truncated protein in increasing insecticidal activity is known in the art. See for example Koziel et al., 1993, Field performance of elite transgenic maize plants expressing an insecticidal protein derived from Bacillus thuringiensis. Bio/technology, 11(2), 194-200; teaches expression of insecticidal proteins from Bacillus thuringiensis have been difficult when the native genes from B. thuringiensis were used requiring the use of truncated version of the native lepidopteran active genes for measurable protein and insecticidal activity in the transgenic plant. Koziel further teaches use of a native δ-endotoxin coding region, which has a high A-T content, appears to lead to abnormally low gene expression in plants. Koziel teaches plants in general have a higher G-C content than that found in the δ-endotoxins (page 194, left paragraph 2). Koziel teaches modifying the coding sequence to increase the G-C content of the native gene results in a dramatic increase in expression of the insecticidal protein (page 194, right paragraph 1). Koziel teaches about field performance of maize plant containing synthetic gene encoding truncated version of the CryIA(b) protein derived from B. thuringiensis which was affected by use of CaMV 35S promoter from maize where the transgenic plant produced higher levels of insecticidal protein (page 194, right paragraph 2).
Furthermore, Deist et al., 2014, Bt toxin modification for enhanced efficacy. Toxins, 6(10), 3005-3027; teaches truncation of Bt toxins has proven to be a useful option for enhancing Bt toxin activity by circumventing the toxin activation step resulting in improved toxicity against target pests and such toxin and truncation also improves expression levels in planta (page 3008, paragraph 4). Deist teaches higher insecticidal activity of truncated Cry1A over Cry1A protoxin for Mandusca sexta and it was not toxic to the plants (page 3009, paragraph 5). Deist further provides Table 1 (page 3018) which shows truncation as a modification for improved efficiency citing various references for different Bt toxin and different insect species. Hence applicant’s arguments are not persuasive.
Furthermore, the issue with Dr. Hjelle declaration is comparing activity of the claimed fragment to activity of the full-length protein taught by Baum et al., but the rationale of the Rejection was directed to obviousness for using the fragment taught by Aziz et al. Therefore, unless the declaration compares to the fragment taught by Aziz, then it is not addressing obviousness rejection at all. Furthermore, the toxicity is happening with the truncated SEQ ID NO: 3 which Aziz found to have insecticidal property. The codon optimization has nothing to do with toxicity it is to do with expression in an organism.
Since codon optimization is routine procedure to express a gene across different species and it would produce the same protein disclosed by Aziz et al. it is not an inventive step. Furthermore, Puigbo teaches about the web server utility that optimizes a DNA or protein sequence. Puigbo further teaches that "optimizer" software can be used to design any new genes that confer new metabolic capabilities in a given species (Page W130, first paragraph).
Response to Declaration of Dr. Ron Lirette
Declaration of Dr. Lirette argues it is known that optimization of codons for heterologous protein expression can result in improved production based on preferred codon utilization in the target species. Dr. Lirette argues there are many factors that influence optimal codons (e.g., size of pools of transfer RNA, avoidance of targeted degradation of “foreign” sequences in the recipient species, etc.), and the choice of which codons to choose is a complex issue. Dr. Lirette argues consider there are 61 codons that encode 20 amino acids, meaning that there is tremendous variability possible at the nucleotide level that can result in production of the same protein; each amino acid can have between 1 and 6 possible codons, so a protein that is 625 ammo acids in length has an astronomical number of combinations that yield the same protein sequence. Dr. Lirette argues for example, a 20 amino acid protein with an average of 4 possible codons for each position has over a trillion possible combinations of codons yielding the same 20 amino acid sequence (Dr. Lirette Declaration, page 2, second paragraph).
Dr. Lirette asserts tools exist to optimize sequences for various species. Dr. Lirette argues the output of each of the tools is influenced by the algorithms used by each, as well as the data sets used to train the tools. Dr. Lirette argues the outputs are variable in terms of recommended codon sequences, and the assessment as to which of the proposed sequences or combinations of sequences will work in practice or will provide the desired result.
Dr. Lirette argues it should be emphasized that “Optimizer" in only one of several tools for codon optimization that produce quite different outputs for the same input and there is no teaching on cited art as whether a given codon optimization software would provide preferable results over competing options. Dr. Lirette argues even the '"Optimizer"' software of Puigbo produces quite different outputs for the same input if different parameters are chosen and there is no clear guidance on the art regarding which parameter would produce preferable results. Dr. Lirette argues for example the nucleotide sequence of the Cry1Da protein that is the subject of the patent application was used as a test for several publicly available optimization programs and to determine concordance among the outputs; different training sets were used among the different algorithms. Dr. Lirette argues Optimizer 1-5 are all the same program, but each of the 5 is based on a different training set. Dr. Lirette argues table below shows results of optimization. "% Identity" and "Identities'"' refer to values obtained compared to the patent sequence. Dr. Lirette argues there is clear variability among the models, even when the same program and training set were used in the analysis. (i.e., Optimizer 1 ). Dr. Lirette argues the choice of which sequence to utilize is not obvious based solely on the algorithms, instead relying on the expertise and judgment of the researcher, \Which is in fact the art underpinning the present patent application (Dr. Lirette Declaration, page3, first paragraph).
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Dr. Lirette argues as demonstrated above, using "Optimizer" software of Puigbo to optim1ze the native Cry 1Da nucleotide sequence for maize does not result in the claimed sequence of SEQ ID NO: 1. Dr. Lirette argues Even the closest parameter still differs by one nucleotide sequence (Dr. Lirette Declaration, page4, first paragraph).
Dr. Lirette argues even if a person skilled in the art would choose ""optimizer" software amongst many possible alternatives, following the disclosure of Puigbo, a person skilled in the art seeking to optimize a truncated version of the native Bt nucleotide sequence of Baum would not arrive at SEQIDNO: 1 (Dr. Lirette Declaration, page4, second paragraph).
Dr. Lirette argues Merlo & Folks does not overcome the deficiencies of Puigbo. The opposite is true for the following reasons:
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Arguments of Dr. Lirette’s declaration were fully considered but they are not found persuasive since:
Regarding argument on improved production, increasing production of a known protein disclosed by Aziz et al. it was not found persuasive and it is not an inventive step. The effect as insecticide is caused by the property of the protein and not by higher production. Furthermore, the invention is directed to the insecticidal activity of the protein not increased production of the protein in the plant.
Since codon optimization is routine procedure to express a gene across different species and it would produce the same protein disclosed by Aziz et al. it is not an inventive step. Furthermore, Puigbo teaches about the web server utility that optimizes a DNA or protein sequence. Puigbo further teaches that "optimizer" software can be used to design any new genes that confer new metabolic capabilities in a given species (Page W130, first paragraph).
The codon optimization is a routine procedure to produce the already known protein taught by Aziz et al. as having insecticidal activity. The claim's patentability hinges on the novelty of the product itself, not the process. The feely available website accessed at http://genomes.urv.es/OPTIMIZER/, accessed at 09/04/2025 as taught by Puigbo et al. showed optimizer is an on-line PHP application that optimizes the codon usage of a DNA sequence to increase its expression level. Therefore, increase in expression is expected. Furthermore, the codon usage database showed it clearly provides codon usage for the maize, for example see the usage table showed below accessed from the linked codon usage database in website.
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Furthermore, Spec, Paragraph 0122 teaches that the definition of the sequence for synthesizing the Bt gene present in the gene construct codes for a 625-amino acid protein corresponding to cry1 Da protein active site and was based on three aspects: (i) Presence of C-terminal domain (Endotoxin C), Central domain (Endotoxin M) and N-terminal domain (Endotoxin N); (ii) Size of the protein active core - residues from aa 1 to 625 (Abdul Aziz, H., Wei Hong, L. and Yusoff, K. Comparative study of cry1 D gene expressed in E. coli and Baculovirus expression system / http://www.ncbi.nlm.nih.gov/protein/AFK29089.1 ); (iii) In silica detection of cry1 Da protoxin hydrolysis sites with trypsin or chymotrypsin enzymes. For this reason, all the aspects of the invention were disclosed by Aziz et al. including the active core, C, M and N terminals of the protein.
Since the Aziz et al. had disclosed the protein locus AFK29089.1 and the coding sequence JQ910166.1 comprises active core crystal toxin protein ID and Baum further teaches their disclosed insecticidal proteins for example SEQ ID NO: 2 can be in truncated forms wherein one or more amino acids are deleted from the N-terminal end, C-terminal end, the middle of the protein, or combinations thereof without a loss of insect inhibitory activity and these fragments should retain the insect inhibitory activity of the parent engineered insecticidal protein (page 7, paragraph 0080). Someone skilled in the art would search for the truncated protein of Baum et al. with the active core crystal toxin protein 1D and would lead to the truncated protein disclosed by Aziz et al. with predictable result of toxicity to some insect pests.
In KSR International Co. v. Teleflex Inc., 550 U.S. 398, 82 USPQ2d 1385 (2007), the Supreme Court held that “obvious to try” was a valid rationale for an obviousness finding, for example, when there is a “design need” or “market demand” and there are a “finite number” of solutions. 550 U.S. at 421, 82 USPQ2d at 1397 (“The same constricted analysis led the Court of Appeals to conclude, in error, that a patent claim cannot be proved obvious merely by showing that the combination of elements was ‘[o]bvious to try.’ ... When there is a design need or market pressure to solve a problem and there are a finite number of identified, predictable solutions, a person of ordinary skill has good reason to pursue the known options within his or her technical grasp.
Furthermore, Aziz discloses the information is dated 03/2018 before the effective date of filling of the invention in https://www.researchgate.net/publication/323627747_Bacillus_thuringiensis_strain_aizawai_active_core_crystal_toxin_protein_1D_gene_partial_cds_ACCESSION_JQ910166, accessed 03/07/2025 that their data showed the active core cry1D gene sequence from genomic DNA of Bacillus thuringiensis subsp. Aizawai and the gene coded for the active form of Cry1D crystal protein and has been proven toxic to its specific targeted-insect pests which were Spodoptera litura and Spodoptera exigua (see previously send OA for enclosed snippet and the entered accession JQ910166.1 included with the website). The attached locus of the ResearchGate website JQ910166.1 encodes SEQ ID NO:3. For this reason it was known to have been effect to insects from genus Spodoptera.
Therefore, the arguments are not persuasive and 103 rejection is maintained.
However, Applicant’s reply is considered to be a bona fide attempt at a response and is being accepted as a complete response.
Claim Rejections - 35 USC § 112 - Indefiniteness
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim 6 is rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 6 depends on cancelled claim 5, therefore the metes and bound of the recited construct cannot be determined.
The claim will be analyzed assuming it would depend on the construct of claim 4.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Obvious over Baum and further in view of Aziz, Merlo and Folkers, Puigbo, and Pardo-Lopez.
Claims 1, 4, 7 and 11-29 are rejected under 35 U.S.C. 103 as being unpatentable over Baum et al. (US Patent publication number: US 2016/0108426A1, published on 04/21/2016); and further in view of Aziz et al. (2012),Comparative study of cry1D gene expressed in E. coli and Baculovirus expression system; Submitted (09-APR-2012); Department of Biology, Universiti Pendidikan Sultan Idris, Faculty of Science and Mathematics, Tanjong Malim, Perak 35900, Malaysia, further in view of, Merlo and Folkerts (US Patent number: 6,166,302, published on 12/26/2000), further in view of Puigbo et al. 2007.OPTIMIZER: A web server utility that optimize a DNA or Protein sequence. Nucleic Acids Research, 35:W126-W131 (see IDS filed on 03/15/2023), and further in view of Pardo-Lopez et al., 2009, Strategies to improve the insecticidal activity of Cry toxins from Bacillus thuringiensis. Peptides, 30(3), 589-595.
The claims are drawn to a codon optimized nucleic acid molecule consisting of SEQ ID NO:1 and encoding a CRY1DA consisting of SEQ ID NO: 3. Claims are further drawn to a nucleic acid construct comprising the nucleic acid molecule. A vector, a host cell, a plant cell, a transgenic plant, comprising the nucleic acid molecule. The claims are further drawn to a cell transformation method comprising introducing into the cell the nucleic acid molecule wherein the nucleic acid molecule integrates into the cell genome and regenerating the transgenic plant from the plant cell. The claims are drawn to a method of producing a transgenic plant comprising transforming the plant cell with the nucleic acid molecule and selecting the plant cell transformed with the nucleic acid molecule. The claims are further drawn to wherein the transgenic plant, a monocotyledonous plant for example maize, rice, sorghum etc., is resistant to crop pests, an insect of the order Lepidoptera and species Spodoptera frugiperda and Diatrea saccharalis. The claims are further drawn to a method of controlling invertebrate pests in crop plants comprising the nucleic acid molecule which comprises planting seeds obtained from the plant in an area of cultivation of crop plants susceptible to invertebrate pests.
Regarding claim 1, Baum et al. teaches about synthesis of genes and their derivatives encoding engineered insecticidal protein and scaffold protein for expression in plant by avoiding ATTA and A/T rich plant sequences and preserving original amino acid sequences (Paragraph 0120 and 0121, Example 4). Instant application uses nucleotide sequence, SEQ ID NO: 2 which is originally isolated from B. thurengiensis to optimize the codon in Optimizer software. The SEQ ID NO: 2 is 99.8% identical to SEQ ID NO: 1 of instant application (see alignment below) which is nucleotide sequence encoding a Cry1Da1 protein taught by Baum et al. (Page 3, Paragraph 0023) suggesting both instant application and Baum et al. started optimizing same native sequences wherein instant application optimized known truncated sequence of Aziz et al. Baum further teaches their disclosed insecticidal proteins for example SEQ ID NO: 2 can be in truncated forms wherein one or more amino acids are deleted from the N-terminal end, C-terminal end, the middle of the protein, or combinations thereof without a loss of insect inhibitory activity and these fragments should retain the insect inhibitory activity of the parent engineered insecticidal protein (page 7, paragraph 0080).
Baum et al. does not teach codon optimized nucleic acid molecule encoding Cry1Da protein that consist of SEQ ID NO: 3 which is a truncated version of Baum’s SEQ ID NO: 2.
Furthermore, it was known in the art before the effective date of filing invention that the truncation of protein can improve the insecticidal activity of Cry toxins from Bacillus thuringiensis. Pardo-Lopez et al. teaches deletion of small fragments from amino-terminal region led to improved toxicity or overcome resistance representing interesting alternatives for insect pest control (page 589, Abstract and page 593).
Aziz et al. teaches a locus AFK29089 which has amino acid sequence which has 100% sequence identity to SEQ ID NO: 3 of the instant application where both has same size of 625 amino acid sequences as truncated protein (see alignment below). Aziz et al. define the locus as an active core crystal toxin protein 1D, from B. thuringiensis (see DEFINITION). Furthermore Aziz et al. teaches the sequence has been disclosed in the comparative study of cry1D gene expressed in E. coli and Baculovirus expression system (see TITLE). Aziz et al. teaches isolation source of the protein is commercial Bt based insecticide. Aziz et al. teaches region from amino acid sequence position 48-250 include a delta endotoxin, N-terminal domain (Endotoxin N), and 460-592, a C-terminal domain (Endotoxin C) and 258-450, an Endotoxin M domain (see the description of the locus below).
Applicant describes Bt gene present in construct were synthesized based on: presence of C-terminal (Endotoxin C), central domain (Endotoxin M) and N-terminal domain (Endotoxin N), size of the protein active core along with In silico detection of cry1Da protoxin hydrolysis site with trypsin or chymotrypsin enzymes (Spec, page 33-34, paragraph 0122). Applicant describes codon of the active core was modified – residues from aa 1-625 /1875 bp nucleotide originally isolated from B. thuringiensis (SEQ ID NO: 2) (Spec, page 34, paragraph 0123).
However, Baum et al. and Aziz et al. do not expressly teach about codon optimizing initially isolated strain of BT 1132C with CG content of 37.91% to a CG content of 63.8% in SEQ ID NO: 1 to make it compatible with maize (Spec, Page 34, Example 2, Paragraph 0123).
Further, in view codon optimization to GC content of 63.8% is known in the art. Merlo and Folkerts et al. teaches about optimizing another B. thuringiensis (BT) gene, Cry1A(c) and its truncated versions (Col. 23, lines 14-30), and teaches optimizing by changing of native codons with CG content of 37% to the maize genes of the GC content range of 45-75% where it teaches the range for expressing structural proteins is 48.6-70.5% with mean of 63.6% (Col 8, line 42-47, Table 1). Their optimized codons were 71% homologous to native BT nucleotide sequence encoding insecticidal plant protein (Col. 2, line 35-37) showing substantial changes. They further described the method they disclosed enables one skilled in the art to modify gene(s) that are foreign to a particular plant so that genes are optimally expressed in plants (Col 3, line 22-26).
Baum et al. does not teach about optimizing codon using freely available software “Optimizer”. Puigbo et al. teaches about the web server utility that optimizes a DNA or protein sequence. Puigbo et al. further teaches that "optimizer" software can be used to design any new genes that confer new metabolic capabilities in a given species (Page W130, first paragraph).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to optimize or adapt nucleotide constitution of CRY1DA gene taught by Baum et al. for content of GC and AT that is suitable for expression of protein in plants with teaching, suggestion, or motivation from prior arts (Aziz et al., Merlo and Folkerts et al. and Puigbo et al.). It would have led one of the ordinary skill to optimize the sequence based on suggestions provided by Baum et al. for sequence encoding native CRY1DA and their engineered derivatives. Pardo-Lopez further motivates to use a truncated protein for improving toxicity or overcome insect resistance as alternatives for insect pest control. Hence someone skilled in the art would use the known truncated protein with disclosed by Aziz et al. with its disclosed N, C and central domain information as active core crystal toxin protein 1D choosing from finite number of truncated proteins with such active core, upload in the freely available software “OPTIMIZER” taught by Puigbo et al. for optimizing codons and optimizing genes conferring to express new metabolic capabilities that can efficiently expresses in plants based on the teaching of most suitable GC content of about 63% suggested by Merlo and Folkerts to obtain the predictable result of codon optimized CRY1DA nucleic acid molecule encoding a Cry1Da protein consisting of SEQ ID NO: 3, comprising a nucleic acid sequence of SEQ ID NO: 1.
Regarding claim 4, Baum et al. teaches the polynucleotide construct that encodes one or more of their engineered insect inhibitory proteins (Paragraph 0003, Line 7-9).
Baum et al. teaches the recombinant DNA constructs where the polynucleotide molecule is operably linked to genetic expression elements such as promoter (Paragraph 0084, line 3-9).
Regarding claim 7, Baum et al. teaches about the construct where a 3' UTR sequence is provided 3' of the coding sequence to facilitate termination of transcription. (Paragraph 0123, line 2-26).
Regarding claim 11, Baum et al. teaches the recombinant DNA construct is operably linked to regulatory sequences (Paragraph 0084, line 3-9).
Regarding claim 12, Baum et al. teaches the recombinant DNA construct which has plant multi-gene expression system, each expressing a different protein (i.e., construct comprising multiple nucleotide sequence for example a nucleotide sequence which is at least 70% identical to SEQ ID NO: 1 and SEQ ID No:10) (Paragraph 0086).
Regarding claim 13, Baum et al. teaches a transformation vector containing expression cassettes comprise a polynucleotide sequence designed for use in plant (Paragraph 0125, Example 6).
Regarding claim 14, Baum et al. teaches the plant or bacterial host cell comprising of optimized polynucleotide of SEQ ID NO:33, 35, 37 and 39 (Page 2, Paragraph 0012) (Page 120, claim 9).
Regarding claim 15, Baum et al. teaches the plant (e.g. corn, cotton) cells comprising of optimized SEQ ID NO:39 expressing engineered insect inhibitory protein Cry1Da1_7.nno (Page 14, Paragraph 0128, Paragraph 0132, claim 16).
Regarding claim 16, Baum et al. teaches the transgenic plant comprising of optimized nucleic acid sequence SEQ ID NO:39 expressing engineered insect inhibitory protein Cry1Da1_7.nno (Page 14, Paragraph 0128, Paragraph 0132, claim 16).
Regarding claim 17, Baum et al. teaches a method of transforming host cell which expresses the optimized polynucleotide encoding the insecticidal protein (Page 14, Example 6, Paragraph 0124, 0125, 0126).
Regarding claim 18, Baum et al. teaches the corn, cotton and soybean plant where the optimized polynucleotide encoding engineered insect inhibitory protein are integrated into genome resulting in increased resistance to lepidopteran insects in insect activity test (Page 14-15, Example 7, 8 and 9, Table 6,7, 8 and 9).
Regarding claim 19, Baum et al. teaches a method of producing transgenic plant which expresses the engineered polynucleotide encoding the insecticidal protein (Page 14, Example 6, Paragraph 0124, 0125, 0126).
Regarding claim 20, Baum et al. teaches method of selecting transformed events expressing engineered protein Cry1Da1_7.nno (Page 14, Paragraph 0133; Page 13, Paragraph 0136).
Regarding claim 21, Baum et al. teaches a method of making transgenic plants that comprised the engineered polynucleotide encoding insecticidal protein, where plant are derived from the plant cell by regeneration. (Page 8, Paragraph 0089).
Regarding claim 22, Baum et al. teaches crops (e.g. corn, cotton and soybean) where the engineered polynucleotide are integrated into genome resulting in increased resistance to lepidopteran insects (Page 14-15, Example 7, 8 and 9, Table 6,7, 8 and 9).
Regarding claim 23, Baum et al. teaches method of producing host cell comprising the engineered polynucleotide where host cell includes monocots (Page 2, Paragraph 0012). Baum et al further give example of method of generation of transgenic plant of maize (i.e., corn) which is monocot (Page14, Example 7).
Regarding claim 24, Baum et al. teaches method of generation of transgenic plant of maize which is monocot (Page14, Example 7) and further give example for method can be carried out in sugarcane (Paragraph 0007), sorghum, wheat or grass (i.e., Brachiaria) (Paragraph 0088).
Regarding claim 25, Baum et al. teaches method produces the transgenic plants resistant to insect crop pests (Page 14-15, Example 7, 8 and 9).
Regarding claim 26, Baum et al. teaches method produces the transgenic plants resistant to insect pests from order Lepidoptera (Page 14-15, Example 7, 8 and 9).
Regarding claim 27, Baum et al. teaches the encoded protein are active against Spodoptera frugiperda (Page 5, Paragraph 0073).
Regarding claim 28, Baum et al. teaches the encoded protein are active against Diatrea saccharalis. (Page 5, Paragraph 0073).
Regarding claim 29, Baum et al. teaches a method of producing a seed comprising optimized polynucleotide encoding engineered insecticidal protein, the method comprise planting seeds (Page 3, Paragraph 0017). Baum et al. discloses the method for controlling a Lepidopteran insect, comprising exposing the pest to the transgenic plant (Page 2, Paragraph 0016).
Obvious over Baum and further in view of Aziz, Gelvin, Merlo and Folkerts, Puigbo
Claims 1 and 4, 6-10 are rejected under 35 U.S.C. 103 as being unpatentable over Baum et al., and further in view of Aziz et al., further in view of Gelvin et al. (US Patent publication number: US 2004/0152197A1, Application number: US/14884432, published in 08/05/2004)), further in view of Merlo and Folkerts; further in view of Puigbo et al.
Claims are drawn to a codon optimized CRY1DA nucleic acid molecule comprising a nucleic acid molecule defined as SEQ ID NO.1 which encodes a Cry1Da protein consisting of SEQ ID NO: 3. Claims are further drawn to a nucleic acid construct that comprises of a maize ubiquitin gene promotor sequence, a 3’ UTR terminator sequence called nopaline synthase gene terminator sequence, a selection gene, a duplicated CaMV 35S gene promotor sequence and Tvsp gene terminator.
Regarding claim 4, Baum et al. teaches the polynucleotide construct that encodes one or more of the engineered insect inhibitory proteins (Paragraph 0003, Line 7-9).
Baum et al. teaches the recombinant DNA constructs where the polynucleotide molecule is operably linked to genetic expression elements such as promoter (Paragraph 0084, line 3-9).
Regarding claim 6, Baum et al. teaches about the recombinant DNA constructs where the polynucleotide molecule is operably linked to genetic expression elements such as promoter (Paragraph 0084, line 3-9).
Baum et al. does not expressly teach that the promotor sequence is maize ubiquitin gene promotor sequence.
Further in view, Gelvin et al. teaches about using maize ubiquitin promoter for expression of genes in recombinant construct (Page 9, paragraph 0098). It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to use maize ubiquitin promoter in their construct taught by Gelvin et al. to obtain predictable result of enhanced plant cell transformation optimized for plants.
Regarding claim 7, Baum et al. teaches about the construct where a 3' UTR sequence is provided 3' of the coding sequence to facilitate termination of transcription. (Paragraph 0123, line 2-26).
Regarding claim 8, Baum et al. teaches the construct comprises transcriptional termination sequences (Paragraph 0123).
Baum et al. does not expressly teach that the terminator sequence is nopaline synthase gene terminator sequence.
Gelvin et al. teaches about a recombinant construct which include nopaline synthase terminator, a 3'NOS fused to the 3' region (Page 5, Paragraph 0069) for enhanced plant cell transformation).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to use nopaline synthase terminator in their construct taught by Gelvin et al. to obtain predictable result of enhanced plant cell transformation optimized for plants.
Regarding claim 9, Baum et al. teaches the construct comprises multiple promoter element, enhancer elements or other expression elements known to those of ordinary skill in the art operably linked to boost the expression of the transgenes and terminator sequences (Paragraph 0123). It further teaches for making transgenic plants requires selecting a plant derived from the plant cell that expresses an insect or lepidoptera - inhibitory amount of engineered insecticidal protein (Paragraph 0089, line 3-11).
Baum et al. does not expressly teach that the construct comprises a selection gene. Gelvin et al. teaches a kanamycin resistance gene (Page 5, Paragraph 0068) and hygromycin resistance gene (Page 5, paragraph 0069) for plant selection linked to the recombinant construct. It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to use selection genes taught by Gelvin et al. to obtain predictable result of efficiently selecting the integrated transgene.
Regarding claim 10, Baum et al. teaches the construct comprises multiple promoter elements (Paragraph 0123).
Baum et al. does not expressly teach that the construct comprises duplicated CaMV 35S gene promoter sequence and Tvsp gene terminator sequence.
Gelvin et al. teaches about the CaMV 35S promotor (double promotor region) in the construct to drive the screenable markers and reporter gene. (Page 9, paragraph 0103). Gelvin et al. further teaches the Tvsp gene terminator which encodes for soybean vegetative storage protein is included in the recombinant construct (Page 10, paragraph 0103, Page 4, Paragraph 0058, FIG. 10).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to use duplicated CaMV 35S gene promoter sequence and Tvsp gene terminator sequence taught by Gelvin et al. to obtain predictable result of enhanced plant cell transformation optimized for plants.
Sequence alignment with SEQ ID NO: 3 to Uniprot database:
I3RS06_BACTU
ID I3RS06_BACTU Unreviewed; 625 AA.
AC I3RS06;
DT 05-SEP-2012, integrated into UniProtKB/TrEMBL.
DT 05-SEP-2012, sequence version 1.
DT 25-MAY-2022, entry version 34.
DE RecName: Full=Crystaline entomocidal protoxin {ECO:0000256|ARBA:ARBA00029653};
DE Flags: Fragment;
OS Bacillus thuringiensis.
OC Bacteria; Firmicutes; Bacilli; Bacillales; Bacillaceae; Bacillus;
OC Bacillus cereus group.
OX NCBI_TaxID=1428 {ECO:0000313|EMBL:AFK29089.1};
RN [1] {ECO:0000313|EMBL:AFK29089.1}
RP NUCLEOTIDE SEQUENCE.
RC STRAIN=Aizawai {ECO:0000313|EMBL:AFK29089.1};
RA Abdul Aziz H., Wei Hong L., Yusoff K.;
RT "Comparative study of cry1D gene expressed in E. coli and Baculovirus
RT expression system.";
RL Submitted (APR-2012) to the EMBL/GenBank/DDBJ databases.
CC -!- SIMILARITY: Belongs to the delta endotoxin family.
CC {ECO:0000256|ARBA:ARBA00007819}.
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DR EMBL; JQ910166; AFK29089.1; -; Genomic_DNA.
DR SMR; I3RS06; -.
DR GO; GO:0005102; F:signaling receptor binding; IEA:InterPro.
DR GO; GO:0090729; F:toxin activity; IEA:UniProtKB-KW.
DR GO; GO:0001897; P:cytolysis by symbiont of host cells; IEA:InterPro.
DR GO; GO:0030435; P:sporulation resulting in formation of a cellular spore; IEA:UniProtKB-KW.
DR Gene3D; 1.20.190.10; -; 1.
DR Gene3D; 2.100.10.10; -; 1.
DR InterPro; IPR008979; Galactose-bd-like_sf.
DR InterPro; IPR038979; Pest_crys.
DR InterPro; IPR005638; Pest_crys_C.
DR InterPro; IPR005639; Pest_crys_N.
DR InterPro; IPR036716; Pest_crys_N_sf.
DR InterPro; IPR001178; Pest_cryst_cen_dom.
DR InterPro; IPR036399; Pest_cryst_cen_dom_sf.
DR PANTHER; PTHR37003; PTHR37003; 1.
DR Pfam; PF03944; Endotoxin_C; 1.
DR Pfam; PF00555; Endotoxin_M; 1.
DR Pfam; PF03945; Endotoxin_N; 1.
DR SUPFAM; SSF49785; SSF49785; 1.
DR SUPFAM; SSF51096; SSF51096; 1.
DR SUPFAM; SSF56849; SSF56849; 1.
PE 3: Inferred from homology;
KW Sporulation {ECO:0000256|ARBA:ARBA00022969};
KW Toxin {ECO:0000256|ARBA:ARBA00022656};
KW Virulence {ECO:0000256|ARBA:ARBA00023026}.
FT DOMAIN 48..250
FT /note="Endotoxin_N"
FT /evidence="ECO:0000259|Pfam:PF03945"
FT DOMAIN 258..450
FT /note="Endotoxin_M"
FT /evidence="ECO:0000259|Pfam:PF00555"
FT DOMAIN 460..592
FT /note="Endotoxin_C"
FT /evidence="ECO:0000259|Pfam:PF03944"
FT NON_TER 625
FT /evidence="ECO:0000313|EMBL:AFK29089.1"
SQ SEQUENCE 625 AA; 70313 MW; 1408C676E64E4C3D CRC64;
Query Match 100.0%; Score 3242; DB 106; Length 625;
Best Local Similarity 100.0%;
Matches 625; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 1 MEINNQNQCVPYNCLSNPKEIILGEERLETGNTVADISLGLINFLYSNFVPGGGFIVGLL 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 MEINNQNQCVPYNCLSNPKEIILGEERLETGNTVADISLGLINFLYSNFVPGGGFIVGLL 60
Qy 61 ELIWGFIGPSQWDIFLAQIEQLISQRIEEFARNQAISRLEGLSNLYKVYVRAFSDWEKDP 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 ELIWGFIGPSQWDIFLAQIEQLISQRIEEFARNQAISRLEGLSNLYKVYVRAFSDWEKDP 120
Qy 121 TNPALREEMRIQFNDMNSALITAIPLFRVQNYEVALLSVYVQAANLHLSILRDVSVFGER 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 TNPALREEMRIQFNDMNSALITAIPLFRVQNYEVALLSVYVQAANLHLSILRDVSVFGER 180
Qy 181 WGYDTATINNRYSDLTSLIHVYTNHCVDTYNQGLRRLEGRFLSDWIVYNRFRRQLTISVL 240
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 WGYDTATINNRYSDLTSLIHVYTNHCVDTYNQGLRRLEGRFLSDWIVYNRFRRQLTISVL 240
Qy 241 DIVAFFPNYDIRTYPIQTATQLTREVYLDLPFINENLSPAASYPTFSAAESAIIRSPHLV 300
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 241 DIVAFFPNYDIRTYPIQTATQLTREVYLDLPFINENLSPAASYPTFSAAESAIIRSPHLV 300
Qy 301 DFLNSFTIYTDSLARYAYWGGHLVNSFRTGTTTNLIRSPLYGREGNTERPVTITASPSVP 360
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 301 DFLNSFTIYTDSLARYAYWGGHLVNSFRTGTTTNLIRSPLYGREGNTERPVTITASPSVP 360
Qy 361 IFRTLSYITGLDNSNPVAGIEGVEFQNTISRSIYRKSGPIDSFSELPPQDASVSPAIGYS 420
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 361 IFRTLSYITGLDNSNPVAGIEGVEFQNTISRSIYRKSGPIDSFSELPPQDASVSPAIGYS 420
Qy 421 HRLCHATFLERISGPRIAGTVFSWTHRSASPTNEVSPSRITQIPWVKAHTLASGASVIKG 480
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 421 HRLCHATFLERISGPRIAGTVFSWTHRSASPTNEVSPSRITQIPWVKAHTLASGASVIKG 480
Qy 481 PGFTGGDILTRNSMGELGTLRVTFTGRLPQSYYIRFRYASVANRSGTFRYSQPPSYGISF 540
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 481 PGFTGGDILTRNSMGELGTLRVTFTGRLPQSYYIRFRYASVANRSGTFRYSQPPSYGISF 540
Qy 541 PKTMDAGEPLTSRSFAHTTLFTPITFSRAQEEFDLYIQSGVYIDRIEFIPVTATFEAEYD 600
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 541 PKTMDAGEPLTSRSFAHTTLFTPITFSRAQEEFDLYIQSGVYIDRIEFIPVTATFEAEYD 600
Qy 601 LERAQKVVNALFTSTNQLGLKTDVT 625
|||||||||||||||||||||||||
Db 601 LERAQKVVNALFTSTNQLGLKTDVT 625
Sequence alignment to SEQ ID NO: 1 to published application database:
US-14-884-432-1; Sequence 1, Application US/14884432
; Patent No. 10059959
; GENERAL INFORMATION
; APPLICANT: Monsanto Technology LLC
; APPLICANT:Baum, James A.
; APPLICANT:Cerruti, Thomas
; APPLICANT:Flasinski, Stanislaw
; APPLICANT:Fu, Xiaoran
; APPLICANT:Howe, Arlene R.
; APPLICANT:Salvador, Sara
; TITLE OF INVENTION: Lepidopteran-Active Cry1Da1 Amino Acid Sequence Variant Proteins
; FILE REFERENCE: P34223US02
; CURRENT APPLICATION NUMBER: US/14/884,432
; CURRENT FILING DATE: 2015-10-15
; PRIOR APPLICATION NUMBER: US 62/064,994
; PRIOR FILING DATE: 2014-10-16
; PRIOR APPLICATION NUMBER: US 62/065,017
; PRIOR FILING DATE: 2014-10-17
; NUMBER OF SEQ ID NOS: 44
; SOFTWARE: PatentIn version 3.5
; SEQ ID NO 1
; LENGTH: 3498
; TYPE: DNA
; ORGANISM: Bacillus thuringiensis
; FEATURE:
; NAME/KEY: misc_feature
; LOCATION: (1)..(3498)
; OTHER INFORMATION: Nucleotide sequence used for expression in a bacterial cell
; OTHER INFORMATION:encoding Cry1Da1.
US-14-884-432-1
Query Match 99.8%; Score 1873.8; DB 1; Length 3498;
Best Local Similarity 99.9%;
Matches 1875; Conservative 0; Mismatches 2; Indels 0; Gaps 0;
Qy 1 ATGGAAATAAATAATCAAAACCAATGTGTGCCTTACAATTGTTTAAGTAATCCTAAGGAG 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 ATGGAAATAAATAATCAAAACCAATGTGTGCCTTACAATTGTTTAAGTAATCCTAAGGAG 60
Qy 61 ATAATATTAGGCGAGGAAAGGCTAGAAACAGGGAATACTGTAGCAGACATTTCATTAGGG 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 ATAATATTAGGCGAGGAAAGGCTAGAAACAGGGAATACTGTAGCAGACATTTCATTAGGG 120
Qy 121 CTTATTAATTTTCTATATTCTAATTTTGTACCAGGAGGAGGATTTATAGTAGGTTTACTA 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 CTTATTAATTTTCTATATTCTAATTTTGTACCAGGAGGAGGATTTATAGTAGGTTTACTA 180
Qy 181 GAATTAATATGGGGATTTATAGGGCCTTCGCAATGGGATATTTTTTTAGCTCAAATTGAG 240
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 GAATTAATATGGGGATTTATAGGGCCTTCGCAATGGGATATTTTTTTAGCTCAAATTGAG 240
Qy 241 CAATTGATTAGTCAAAGAATAGAAGAATTTGCTAGGAATCAGGCAATTTCAAGATTGGAG 300
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 241 CAATTGATTAGTCAAAGAATAGAAGAATTTGCTAGGAATCAGGCAATTTCAAGATTGGAG 300
Qy 301 GGGCTAAGCAATCTTTATAAGGTCTATGTTAGAGCGTTTAGCGACTGGGAGAAAGATCCT 360
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 301 GGGCTAAGCAATCTTTATAAGGTCTATGTTAGAGCGTTTAGCGACTGGGAGAAAGATCCT 360
Qy 361 ACTAATCCTGCTTTAAGGGAAGAAATGCGTATACAATTTAATGACATGAATAGTGCTCTC 420
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 361 ACTAATCCTGCTTTAAGGGAAGAAATGCGTATACAATTTAATGACATGAATAGTGCTCTC 420
Qy 421 ATAACGGCTATTCCACTTTTTAGAGTTCAAAATTATGAAGTTGCTCTTTTATCTGTATAT 480
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 421 ATAACGGCTATTCCACTTTTTAGAGTTCAAAATTATGAAGTTGCTCTTTTATCTGTATAT 480
Qy 481 GTTCAAGCCGCAAACTTACATTTATCTATTTTAAGGGATGTTTCAGTTTTCGGAGAAAGA 540
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 481 GTTCAAGCCGCAAACTTACATTTATCTATTTTAAGGGATGTTTCAGTTTTCGGAGAAAGA 540
Qy 541 TGGGGATATGATACAGCGACTATCAATAATCGCTATAGTGATCTGACTAGCCTTATTCAT 600
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 541 TGGGGATATGATACAGCGACTATCAATAATCGCTATAGTGATCTGACTAGCCTTATTCAT 600
Qy 601 GTTTATACTAACCATTGTGTGGATACGTATAATCAGGGATTAAGGCGTTTGGAAGGTCGT 660
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 601 GTTTATACTAACCATTGTGTGGATACGTATAATCAGGGATTAAGGCGTTTGGAAGGTCGT 660
Qy 661 TTTCTTAGCGATTGGATTGTATATAATCGTTTCCGGAGACAATTGACAATTTCAGTATTA 720
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 661 TTTCTTAGCGATTGGATTGTATATAATCGTTTCCGGAGACAATTGACAATTTCAGTATTA 720
Qy 721 GATATTGTTGCGTTTTTTCCAAATTATGATATTAGAACATATCCAATTCAAACAGCTACT 780
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 721 GATATTGTTGCGTTTTTTCCAAATTATGATATTAGAACATATCCAATTCAAACAGCTACT 780
Qy 781 CAGCTAACGAGGGAAGTCTATCTGGATTTACCTTTTATTAATGAAAATCTTTCTCCTGCA 840
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 781 CAGCTAACGAGGGAAGTCTATCTGGATTTACCTTTTATTAATGAAAATCTTTCTCCTGCA 840
Qy 841 GCAAGCTATCCAACCTTTTCAGCTGCTGAAAGTGCTATAATTAGAAGTCCTCATTTAGTA 900
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 841 GCAAGCTATCCAACCTTTTCAGCTGCTGAAAGTGCTATAATTAGAAGTCCTCATTTAGTA 900
Qy 901 GACTTTTTAAATAGCTTTACCATTTATACAGATAGTCTGGCACGTTATGCATATTGGGGA 960
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 901 GACTTTTTAAATAGCTTTACCATTTATACAGATAGTCTGGCACGTTATGCATATTGGGGA 960
Qy 961 GGGCACTTGGTAAATTCTTTCCGCACAGGAACCACTACTAATTTGATAAGATCCCCTTTA 1020
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 961 GGGCACTTGGTAAATTCTTTCCGCACAGGAACCACTACTAATTTGATAAGATCCCCTTTA 1020
Qy 1021 TATGGAAGGGAAGGAAATACAGAGCGCCCCGTAACTATTACCGCATCACCTAGCGTACCA 1080
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1021 TATGGAAGGGAAGGAAATACAGAGCGCCCCGTAACTATTACCGCATCACCTAGCGTACCA 1080
Qy 1081 ATATTTAGAACACTTTCATATATTACAGGCCTTGACAATTCAAATCCTGTAGCTGGAATC 1140
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1081 ATATTTAGAACACTTTCATATATTACAGGCCTTGACAATTCAAATCCTGTAGCTGGAATC 1140
Qy 1141 GAGGGAGTGGAATTCCAAAATACTATAAGTAGAAGTATCTATCGTAAAAGCGGTCCAATA 1200
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1141 GAGGGAGTGGAATTCCAAAATACTATAAGTAGAAGTATCTATCGTAAAAGCGGTCCAATA 1200
Qy 1201 GATTCTTTTAGTGAATTACCACCTCAAGATGCCAGCGTATCTCCTGCAATTGGGTATAGT 1260
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1201 GATTCTTTTAGTGAATTACCACCTCAAGATGCCAGCGTATCTCCTGCAATTGGGTATAGT 1260
Qy 1261 CACCGTTTATGCCATGCAACATTTTTAGAACGGATTAGTGGACCAAGAATAGCAGGCACC 1320
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1261 CACCGTTTATGCCATGCAACATTTTTAGAACGGATTAGTGGACCAAGAATAGCAGGCACC 1320
Qy 1321 GTATTTTCTTGGACACACCGTAGTGCCAGCCCTACTAATGAAGTAAGTCCATCTAGAATT 1380
|||||||||||||||||||||||||||||||||||||| |||||||||||||||||||||
Db 1321 GTATTTTCTTGGACACACCGTAGTGCCAGCCCTACTAACGAAGTAAGTCCATCTAGAATT 1380
Qy 1381 ACACAAATTCCATGGGTAAAGGCGCATACTCTTGCATCTGGTGCCTCCGTCATTAAAGGT 1440
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1381 ACACAAATTCCATGGGTAAAGGCGCATACTCTTGCATCTGGTGCCTCCGTCATTAAAGGT 1440
Qy 1441 CCTGGATTTACAGGTGGAGATATTCTGACTAGGAATAGTATGGGCGAGCTGGGGACCTTA 1500
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1441 CCTGGATTTACAGGTGGAGATATTCTGACTAGGAATAGTATGGGCGAGCTGGGGACCTTA 1500
Qy 1501 CGAGTAACCTTCACAGGAAGATTACCACAAAGTTATTATATACGTTTCCGTTATGCTTCG 1560
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1501 CGAGTAACCTTCACAGGAAGATTACCACAAAGTTATTATATACGTTTCCGTTATGCTTCG 1560
Qy 1561 GTAGCAAATAGGAGTGGTACATTTAGATATTCACAGCCACCTTCGTATGGAATTTCATTT 1620
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1561 GTAGCAAATAGGAGTGGTACATTTAGATATTCACAGCCACCTTCGTATGGAATTTCATTT 1620
Qy 1621 CCAAAAACTATGGACGCAGGTGAACCACTAACATCTCGTTCGTTCGCTCATACAACACTC 1680
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1621 CCAAAAACTATGGACGCAGGTGAACCACTAACATCTCGTTCGTTCGCTCATACAACACTC 1680
Qy 1681 TTCACTCCAATAACCTTTTCACGAGCTCAAGAAGAATTTGATCTATACATCCAATCGGGT 1740
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1681 TTCACTCCAATAACCTTTTCACGAGCTCAAGAAGAATTTGATCTATACATCCAATCGGGT 1740
Qy 1741 GTTTATATAGATCGAATTGAATTTATACCGGTTACTGCAACATTTGAGGCAGAATATGAT 1800
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1741 GTTTATATAGATCGAATTGAATTTATACCGGTTACTGCAACATTTGAGGCAGAATATGAT 1800
Qy 1801 TTAGAAAGAGCGCAAAAGGTGGTGAATGCCCTGTTTACGTCTACAAACCAACTAGGGCTA 1860
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1801 TTAGAAAGAGCGCAAAAGGTGGTGAATGCCCTGTTTACGTCTACAAACCAACTAGGGCTA 1860
Qy 1861 AAAACAGATGTGACGTA 1877
||||||||||||||| |
Db 1861 AAAACAGATGTGACGGA 1877
Sequence alignment with SEQ ID NO: 3 to published application database for full length amino acid sequence.
RESULT 6
US-14-884-432-2
; Sequence 2, Application US/14884432
; Publication No. US20160108426A1
; GENERAL INFORMATION
; APPLICANT: Monsanto Technology LLC
; APPLICANT:Baum, James A.
; APPLICANT:Cerruti, Thomas
; APPLICANT:Flasinski, Stanislaw
; APPLICANT:Fu, Xiaoran
; APPLICANT:Howe, Arlene R.
; APPLICANT:Salvador, Sara
; TITLE OF INVENTION: Lepidopteran-Active Cry1Da1 Amino Acid Sequence Variant Proteins
; FILE REFERENCE: P34223US02
; CURRENT APPLICATION NUMBER: US/14/884,432
; CURRENT FILING DATE: 2015-10-15
; PRIOR APPLICATION NUMBER: US 62/064,994
; PRIOR FILING DATE: 2014-10-16
; PRIOR APPLICATION NUMBER: US 62/065,017
; PRIOR FILING DATE: 2014-10-17
; NUMBER OF SEQ ID NOS: 44
; SOFTWARE: PatentIn version 3.5
; SEQ ID NO 2
; LENGTH: 1165
; TYPE: PRT
; ORGANISM: Bacillus thuringiensis
; FEATURE:
; NAME/KEY: MISC_FEATURE
; LOCATION: (1)..(1165)
; OTHER INFORMATION: Amino acid sequence of the protein Cry1Da1.
US-14-884-432-2
Query Match 100.0%; Score 3242; DB 14; Length 1165;
Best Local Similarity 100.0%;
Matches 625; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 1 MEINNQNQCVPYNCLSNPKEIILGEERLETGNTVADISLGLINFLYSNFVPGGGFIVGLL 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 MEINNQNQCVPYNCLSNPKEIILGEERLETGNTVADISLGLINFLYSNFVPGGGFIVGLL 60
Qy 61 ELIWGFIGPSQWDIFLAQIEQLISQRIEEFARNQAISRLEGLSNLYKVYVRAFSDWEKDP 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 ELIWGFIGPSQWDIFLAQIEQLISQRIEEFARNQAISRLEGLSNLYKVYVRAFSDWEKDP 120
Qy 121 TNPALREEMRIQFNDMNSALITAIPLFRVQNYEVALLSVYVQAANLHLSILRDVSVFGER 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 TNPALREEMRIQFNDMNSALITAIPLFRVQNYEVALLSVYVQAANLHLSILRDVSVFGER 180
Qy 181 WGYDTATINNRYSDLTSLIHVYTNHCVDTYNQGLRRLEGRFLSDWIVYNRFRRQLTISVL 240
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 WGYDTATINNRYSDLTSLIHVYTNHCVDTYNQGLRRLEGRFLSDWIVYNRFRRQLTISVL 240
Qy 241 DIVAFFPNYDIRTYPIQTATQLTREVYLDLPFINENLSPAASYPTFSAAESAIIRSPHLV 300
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 241 DIVAFFPNYDIRTYPIQTATQLTREVYLDLPFINENLSPAASYPTFSAAESAIIRSPHLV 300
Qy 301 DFLNSFTIYTDSLARYAYWGGHLVNSFRTGTTTNLIRSPLYGREGNTERPVTITASPSVP 360
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 301 DFLNSFTIYTDSLARYAYWGGHLVNSFRTGTTTNLIRSPLYGREGNTERPVTITASPSVP 360
Qy 361 IFRTLSYITGLDNSNPVAGIEGVEFQNTISRSIYRKSGPIDSFSELPPQDASVSPAIGYS 420
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 361 IFRTLSYITGLDNSNPVAGIEGVEFQNTISRSIYRKSGPIDSFSELPPQDASVSPAIGYS 420
Qy 421 HRLCHATFLERISGPRIAGTVFSWTHRSASPTNEVSPSRITQIPWVKAHTLASGASVIKG 480
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 421 HRLCHATFLERISGPRIAGTVFSWTHRSASPTNEVSPSRITQIPWVKAHTLASGASVIKG 480
Qy 481 PGFTGGDILTRNSMGELGTLRVTFTGRLPQSYYIRFRYASVANRSGTFRYSQPPSYGISF 540
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 481 PGFTGGDILTRNSMGELGTLRVTFTGRLPQSYYIRFRYASVANRSGTFRYSQPPSYGISF 540
Qy 541 PKTMDAGEPLTSRSFAHTTLFTPITFSRAQEEFDLYIQSGVYIDRIEFIPVTATFEAEYD 600
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 541 PKTMDAGEPLTSRSFAHTTLFTPITFSRAQEEFDLYIQSGVYIDRIEFIPVTATFEAEYD 600
Qy 601 LERAQKVVNALFTSTNQLGLKTDVT 625
|||||||||||||||||||||||||
Db 601 LERAQKVVNALFTSTNQLGLKTDVT 625
Response to Arguments
Applicant's arguments filed 07/14/2025 have been fully considered but they are not persuasive.
Applicant argues present claims are not directed to the Cry1Da protein in and of itself. Applicant argues present claims are directed to: "[a] codon-optimized nucleic acid molecule consisting of the nucleotide sequence of SEQ ID NO: 1;" a nucleic acid construct, vector, a host cell, a plant cell, and a transgenic plant comprising said nucleic acid molecule; and methods using said nucleic acid molecule (Response to rejection, page 9, paragraph 3). Applicant argues the proper question is whether it would have been obvious to produce said codon-optimized nucleic acid molecule and compositions comprising said molecule and to use it in the presently claimed methods (Response to rejection, page 9, second to last paragraph).
Applicant argues in the present claims, the cited references provide no indication that the codon-optimized nucleic acid molecule could be successfully used in pest control and, therefore, provide no motivation to one of ordinary skill in the art to obtain and use such a molecule (Response to rejection, page 10, first paragraph).
Applicant argues on pages 5-6 of the Office Action, the Examiner's asserts that Cry1Da protein of SEQ ID NO: 3 was known to be toxic. Applicant argues in making this assertion, the Examiner appears to rely on the fact that the Definition section for GenBank entry AFK29089. 1 states "active core crystal toxin protein 1D, partial" and on the disclosure of another Aziz reference mentioned in the Office Action on pages 7 and 12 (Response to rejection, page 10, paragraph 2).
Applicant asserts Aziz discloses SEQ ID NO: 3. Applicant argues neither Aziz nor the other cited references provide a reasonable expectation that (1) this particular truncated Cry1Da protein will have increased and commercially useful insecticidal activity, and (2) that optimizing the nucleotide sequence encoding this particular protein would allow one to successfully express it in a plant and achieve said increased insecticidal activity (Response to rejection, page 10, paragraph 3).
Applicant argues the cited references are completely silent with respect to SEQ ID NO: 1 encoding said protein. Applicant argues nothing in the cited references would guide one of skill in the art to select the specific Cry1Da encoding sequence for optimization with any reasonable expectation of success. Applicant argues of course, absolutely nothing in the cited references would have provided any expectations that such surprisingly high insecticidal activity demonstrated by Applicants could be achieved with this sequence. Applicant argues the results demonstrated by Applicants as described in the specification and in the Declaration by Dr. Camilo are certainly probative of non in the specification and in the Declaration by Dr. Camilo are certainly probative of nonobviousness and are, in fact, surprising for reasons set forth in the Declaration and discussed below (Response to rejection, page 10, last paragraph).
Applicant argues Baum teaches that the wild type Cry1Da is not commercially useful. Applicant argues Specifically, Baum states: "Because of its narrow insecticidal spectrum and its inability to provide commercial-level protection against a range of important Lepidopteran agricultural pests such as CEW, the Cry1Da1 insecticidal protein has limited value as a transgenic plant insect control trait." Paragraph [0005] of Baum (Response to rejection, page 11, first paragraph).
Applicant argues Baum also teaches away from using a C-terminally truncated Cry1Da protein, such as the Cry1Da protein consisting of SEQ ID NO: 3 recited in the present claims. Applicant argues Baum teaches using the full-length amino acid sequence of variants of Cry1Dal or a chimeric protein named TIC844, which comprises the Cry1Da1 N-terminal domain and the Cry1Ab3 protoxin C-terminal domain. See paragraph [0008] of Baum. Applicant argues Baum teaches that using said variants, which all require a C-terminal protoxin domain either from Cry1Da1 or Cry1Ab3, would "exhibit markedly improved activity (compared to the Cry1Da1 native toxin) towards H. zea." Paragraph [0007] of Baum (Response to rejection, page 11, second paragraph).
Applicant argues Baum teaches a person skilled in the art to not use the C-terminally truncated sequence of SEQ ID NO: 3. Applicant argues therefore, a person skilled in the art reading Baum would not have been motivated to construct a codon optimized nucleotide sequence encoding a wild-type Cry1Da protein, let alone a C-terminally truncated version thereof recited in the present claims upon in the Declaration of Dr. Hjelle submitted herewith, there is nothing in Aziz or any other Applicant argues as explained in Dr. Camilo' s Declaration, the annotation of most database entries, such as that of Aziz, is made solely based on sequence similarity to known sequences rather than any experimental results (Response to rejection, page 11, last paragraph). Applicant argues there is a very large number of annotations of Cry 1Da proteins with many different truncations, none of which is based on experimental evidence. Applicant argues any annotated sequence could be hypothesized to possibly have insecticidal activity solely based on sequence similarity. Applicant argues one of ordinary skill in the art would have no reasonable expectation of insecticidal activity without any experimental evidence. Applicant argues the Examiner's assertion that selecting the nucleic acid molecule encoding the specific protein sequence of Aziz for codon optimization and expression in a plant for pest control would have been obvious is based on impermissible hindsight (Response to rejection, page 12, first paragraph).
Applicant argues one of ordinary skill in the art would have understood that there is a significant difference between baseline insecticidal activity and commercially useful insecticidal activity. Applicant asserts some insecticidal activity of the Aziz sequence could possibly be hypothesized based solely on sequence similarity, a commercially useful insecticidal activity certainly could not. Applicant argues as addressed in prior Declarations by Dr. Camilo and further expanded upon in the Declaration of Dr. Hjelle submitted herewith, there is nothing in Aziz or any other cited reference that indicates that the specific sequence of Aziz, among many other available options, would have a commercially useful insecticidal activity (Response to rejection, page 12, second paragraph).
Applicant argues Pardo-Lopez teaches that point mutations in Domains I and II in combination with the N-terminal deletion improves the insecticidal activity of the Cry IA toxin. Applicant argues Pardo-Lopez does not teach anything about the toxicity of the C-terminally truncated Cry1Da proteins. Applicant argues Cry1Da protein of SEQ ID NO: 3 is C-terminally truncated in relation to the wild-type sequence disclosed in Baum, rather than N-terminally truncated as taught by Pardo-Lopez (Response to rejection, pages 12 and 13, last and first paragraph).
Applicant argues cited references do not teach the codon-optimized nucleic acid molecule of SEQ ID NO: 1 recited in the present claims and, in fact, teach away from the codon optimization that was performed to obtain the nucleotide sequence of SEQ ID NO: 1. Applicant argues this point is discussed in detail in the Declaration of Dr. Lirette submitted herewith. Applicant argues as explained in Dr. Lirette's Declaration, using "Optimizer" software of Puigbo to optimize the native Bt Cry1Da nucleotide sequence for expression in maize does not result in the sequence of SEQ ID NO: I recited in the present claims (Response to rejection, pages 13, second paragraph).
Applicant argues Merlo does not cure the deficiencies of Puigbo and teaches away from SEQ ID NO:1 (Response to rejection, pages 13).
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Applicant argues based on Puigbo and Merlo, a person skilled in the art seeking to optimize a truncated version of the native Bt nucleotide sequence of Baum would not have arrived at SEQ ID NO:1(Response to rejection, pages 14, second paragraph). Applicant argues Gelvin is cited merely for teaching elements of an expression vector and does not cure the deficiencies of the other references (Response to rejection, pages 14, paragraph 3).
Applicant’s argument were fully considered but they are not found persuasive since:
Regarding argument on Baum et al. teaches away from using a C-terminally truncated Cry1Da protein, Baum further teaches their disclosed insecticidal proteins for example SEQ ID NO: 2 can be in truncated forms wherein one or more amino acids are deleted from the N-terminal end, C-terminal end, the middle of the protein, or combinations thereof without a loss of insect inhibitory activity and these fragments should retain the insect inhibitory activity of the parent engineered insecticidal protein (page 7, paragraph 0080). Furthermore, Applicant is comparing activity of the claimed fragment to activity of the full-length protein taught by Baum et al., but the rationale of the Rejection was directed to obviousness for using the fragment taught by Aziz et al. Therefore, unless the declaration compares to the fragment taught by Aziz, then it is not addressing obviousness rejection at all. Furthermore, the toxicity is happening with the truncated SEQ ID NO: 3 which Aziz found to have insecticidal property. The codon optimization has nothing to do with toxicity it is to do with expression in an organism.
Regarding argument that Aziz teaches any annotated sequence could be hypothesized to possibly have insecticidal activity solely based on sequence similarity. The rationale to modify or combine the prior art of Baum et al. does not have to be expressly stated in the prior art; the rationale may be expressly or impliedly contained in the prior art or it may be reasoned from knowledge generally available to one of ordinary skill in the art, established scientific principles, or legal precedent established by prior case law. In re Fine, 837 F.2d 1071, 5 USPQ2d 1596 (Fed. Cir. 1988); In re Jones, 958 F.2d 347, 21 USPQ2d 1941 (Fed. Cir. 1992); see also In re Kotzab, 217 F.3d 1365, 1370, 55 USPQ2d 1313, 1317 (Fed. Cir. 2000). Furthermore, “Obviousness does not require absolute predictability of success.” Id. at 903, 7 USPQ2d at 1681., but at least some degree of predictability is required. The fact that the Aziz’s locus AFK29089 being truncated protein of Baum et al. and Aziz et al.’s definition as “active core crystal toxin protein 1D” would have scientifically predictable to have insecticidal activity.
Furthermore, Aziz discloses the information is dated 03/2018 before the effective date of filling of the invention in https://www.researchgate.net/publication/323627747_Bacillus_thuringiensis_strain_aizawai_active_core_crystal_toxin_protein_1D_gene_partial_cds_ACCESSION_JQ910166, accessed 03/07/2025 that their data showed the active core cry1D gene sequence from genomic DNA of Bacillus thuringiensis subsp. Aizawai and the gene coded for the active form of Cry1D crystal protein and has been proven toxic to its specific targeted-insect pests which were Spodoptera litura and Spodoptera exigua (see enclosed snippet and the entered accession JQ910166.1 included with the website included in the office action previously presented). The attached locus of the ResearchGate website JQ910166.1 encodes SEQ ID NO:3. For this reason it was known to have effect to insects from genus Spodoptera.
Regarding argument on commercially useful insecticidal activity of Aziz et al.’s locus AFK29089 the strongest rationale for combining references is a recognition, expressly or impliedly in the prior art or drawn from a convincing line of reasoning based on established scientific principles or legal precedent, that some advantage or expected beneficial result would have been produced by their combination. In re Sernaker, 702 F.2d 989, 994-95, 217 USPQ 1, 5-6 (Fed. Cir. 1983).
Regarding argument on cited references do not teach the codon-optimized nucleic acid molecule of SEQ ID NO: 1, the argument is not found persuasive since codon optimization is routine procedure to express a gene across different species and it would produce the same protein disclosed by Aziz et al. it is not an inventive step. Furthermore, Puigbo teaches about the web server utility that optimizes a DNA or protein sequence. Puigbo further teaches that "optimizer" software can be used to design any new genes that confer new metabolic capabilities in a given species (Page W130, first paragraph).
The codon optimization is a routine procedure to produce the already known protein taught by Aziz et al. as having insecticidal activity, although the protein is not produced by optimizing codon. The claim's patentability hinges on the novelty of the product itself, not the process. The feely available website accessed at http://genomes.urv.es/OPTIMIZER/, access at 09/04/2025 as taught by Puigbo et al. showed Optimizer is an on-line PHP application that optimizes the codon usage of a DNA sequence to increase its expression level. Therefore increase in expression is expected. Furthermore, the codon usage database showed it clearly provides codon usage for the maize, for example see the usage table showed below accessed from the linked codon usage database in website.
Therefore, the arguments are not persuasive and 103 rejection is maintained.
However, Applicant’s reply is considered to be a bona fide attempt at a response and is being accepted as a complete response.
Conclusion
No claims are allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Examiner’s Contact Information
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/SANTOSH SHARMA/Examiner, Art Unit 1663
/DAVID H KRUSE/Primary Examiner, Art Unit 1663
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