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 .
Priority
Acknowledgment is made of applicant’s claim for foreign priority of Chinese Application No. CN202310159430.7, filed on February 23, 2023. Receipt is acknowledged of certified copies of papers required by 37 CFR 1.55.
Applicants have not filed a translation of the foreign application CN202310159430.7 to which they are claiming foreign priority.
Claim Interpretation
Based on the disclosure of specification, the cell in claim 5 is interpreted as bacterial cell.
Claim Rejections - 35 USC § 112(b)
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.
Claims 1,2, 5, 6, and 11 are 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 1 recites “encoding gene of domestic pig (Sus scrofa f. domestica)-derived MB (SsMB) sequentially”. There is no special definition of a term “derived” in the specification about a gene that is derived from domestic pig. By using BRI, the term has been interrupted as a region in the genome of domestic pig that is responsible for the myoglobin synthesis. The term is indefinite because the specification does not clearly define the gene necessary for producing domestic pig myoglobin and there exist multiple genes or gene products to which the term "derived" could be referring to. Further, it is unclear to what extent derivatives, variants, and truncations are excluded or included by the claim language. It is established in the art that there exist multiple isoforms of domestic pig myoglobin, and multiple genes which are involved in the production of the myoglobin in pig. For instance, Fernández‐Barroso et al. 2022 (https://doi.org/10.1111/age.13195 ) reported that Iranian pigs with high level of myoglobin content showed higher expression of HBA1 gene indicating that HBA1 gene is responsible for myoglobin production.
Since there are multiple genes and/or different derivatives of the myoglobin that are present in pig, the instant claims are indefinite. Therefore, artisan would not be reasonably apprised of the scope of the claims 1,2, 5, 6, and 11, and therefore, these claims are rejected under 112 (b) for indefiniteness.
Claim Rejections - 35 USC § 103
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 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.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that weqwas not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1 and 5 are rejected under 35 U.S.C. 103 as being unpatentable over United States Pre-Grant Publication (US-PGPUB) No. 20170342132 (“US2017”, published on November 30, 2017).
US2017 teaches a method and compositions for the expression and secretion of heme-containing polypeptides in recombinant bacterial cells such as Bacillus cells or in recombinant plants or plant cells (see abstract and paragraph 0002). US2017 teaches that heme-containing polypeptide can be selected from the group consisting of an androglobin, a cytoglobin, globin E, globin X, globin Y, a hemoglobin, a myoglobin…….(see paragraph 0014). US2017 teaches that culturing such a recombinant bacterium under conditions that allow the heme-containing polypeptide with the signal peptide to be secreted from the bacterium for improved production yield and easier purification of the heme-containing polypeptides (see paragraphs 0006,0012, 0125-0132).
US2017 teaches that heme-containing polypeptide sequence can be found in a mammal including cow, pig, rat, dot, or horse (see paragraph 0039).
US2017 teaches that suitable bacteria for expression of a polypeptide of the disclosure can be gram negative or gram-positive bacteria. For example, the bacteria can be a species of Escherichia (e.g., E. coli) or a species of Bacillus (e.g., B. subtilis) (see paragraph 0116).
US2017 teaches that sequence encoding signal peptide can include phoA, pelB, ompA, and others (see paragraph 0081).
Although US2017 teaches a method and compositions for the expression and secretion of heme-containing polypeptides in recombinant bacterial cells as recited in claims 1 and 5, it does not explicitly teach all the limitations in a single embodiment.
Therefore, it would have been obvious before the effective filing data of the instant inventions to produce an expression vector encoding myoglobin gene from domestic pig and fused with signal peptides because US2017 teaches heme-containing polypeptide, which also includes myoglobin, can be found in pig. Furthermore, US2017 teaches signal peptides fused with heme-containing polynucleotide for the secretion pathway. An artisan would have been motivated by the teachings of US2017 to produce a recombinant vector comprising myoglobin gene and signal peptides because the signal peptides recited in the instant claims are well known and useful peptide for secreting proteins from bacterial cells.
Claims 6 and 11 are rejected under 35 U.S.C. 103 as being unpatentable over US2017 as applied to claims 1 and 5 above, and further in view of Shilling et al. 2020 (Communications Biology, 3:214, “Shilling”).
Teachings of US2017 as applicable to claims 1 and 5 have been described above. However, US2017 does not teach E. coli BL21 (DE3) cell, as recited in claim 6, and the expression vector pET-32a, as recited in claim 11.
Shilling teaches a method of recombinant protein producing in E. coli. Shilling teaches that pET series of expression plasmids are widely used for recombinant protein production (abstract). In particular, Shilling explained about the use of pET28a in protein production. Although Shilling explicitly doesn’t teach pET-32a plasmid vector as claimed in instant claim 11, the authors stated that the design is applicable to most of the 103 vectors in the pET series and can be easily implemented (abstract).
Further, Shilling discusses about the use of transformed BL21(DE3) cells for performing protein expression (see “protein expression and fractionation” section of methods).
Therefore, it would have been obvious for an artisan to clone the cassette of US2017 into a pET series expression vector, such as the claimed pET-32 vector, and introduce the resulting vector into BL21(DE3) cells for recombinant protein expression. An artisan would have been motivated to make this modification because Shilling teaches that its design is applicable to most vector in the pET series and can be easily implemented. An artisan would have had a reasonable expectation that use of the pET-32a vector and BL21(DE3) cells would successfully produce the recombinant protein.
Claims 8 and 9 are rejected under 35 U.S.C. 103 as being unpatentable over US2017 as applied to claims 1 and 5 above, and further in view of Molecular Cloning, A Laboratory Manual by Cold Spring Harbor Laboratory Press, 2001, “Manual”) and Silaban et al. 2019 (IOP Conference Series: Earth and Environmental Science, 217: 012039, “Silaban”).
Teachings of US2017 as applicable to claims 1 and 5 have been described above. However, US2017 does not teach protein expression method, as recited in claim 8 and 9.
Manual teaches expression of genes in E. coli by using IPTG inducible promotors (see chapter 15, protocol 1). The manual teaches about growing the cells in LB medium overnight at the appropriate temperature of 20-37oC for >2 hours in shaking incubator until the cells reach mid-log growth (A550 of 0.5-1) (see method step 5 and 6). The manual also teaches adding isopropyl-3-d-thiogalactoside (IPTG) with a final concentration of 1 mM and continue incubation at 20-37 oC for induction (see method step 8). See below -
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However, manual does not teach about IPTG induction at 180 rpm as recites in claim 8 and IPTG concentration of 0.01 uM, and induction for 12 hours, as recited in claim 9.
Silaban teaches about rhPT-2 gene expression with IPTG induction. Silaban teaches that various concentration of IPTG (0.01 mM, 0.025 mM, 0.05 mM, 0.075 mM, 0.1 mM, 0.2 mM and 0.3 mM) with agitation rate of 100 rpm at a temperature of 12°C was used for induction and cells were harvested after 18 hours of incubation (see “rhPT-2 expression with IPTG induction” under experimental method section). Although Silaban does not explicitly teach induction parameters as claimed in claim 9, these parameters would have been optimized to get the best results and to arrive at the claimed inventions.
Therefore, It would have been obvious for an artisan to modify the IPTG induction parameters of manual with the Silaban because Silaban teaches that IPTG concentration used for gene expression is a critical factor for protein yield and it also has a potential inhibitory effect on cell growth.
Allowable Subject Matter
Claims 2 and 10 are objected to as being dependent upon a rejected claim, but would be allowable if rewritten in independent form including all of the limitations of the base claim and any intervening claims. The subject matter recited in claims 2 and 10 has been found eligible based on prior art search of record because gene expression cassette with the myoglobin gene (SEQ ID NO:4) fused with signal peptides is not disclosed in the art. Based on prior art search, a nucleotide sequence that is about 67.5% identical to SEQ ID NO:4 has been found in Chinese patent application (application no. CN113136349A). Nucleotide sequence in CN113136349A (SEQ ID NO:1) is also a pig myoglobin gene. However, there are multiple differences between the nucleotide sequences (see alignment below), which are due to codon optimization of the claimed sequence.
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There is no teaching of a myoglobin gene as represented by SEQ ID NO:4 combined with signal peptides Pel B and Omp A sequences.
Conclusion
Claims 1, 5-6, 8-9, and 11 are rejected.
Claims 2 and 10 are objected.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to SURAJ SAPKOTA whose telephone number is (571)270-0842. The examiner can normally be reached Monday-Thursday 7am-5pm.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Ram R Shukla can be reached at (571) 272-0735. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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Suraj Sapkota
Patent Examiner
AU 1635
/RAM R SHUKLA/Supervisory Patent Examiner, Art Unit 1635
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