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 .
Election/Restrictions
Applicant’s election of Group II is acknowledge. Claims 1-9, 11-15, 19-22, and 24 are drawn to the elected invention and are examined on the merits. Claims 10, 16-18, and 23 are withdrawn from further consideration pursuant to 47 CFR 1.142(b) as being drawn to a non-elected invention.
Claim Status
Claims 1-24 are pending.
Claims 1-9, 11-15, 19-22, and 24 are examined on the merits.
Claim Rejections - 35 USC § 112
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Scope of Enablement
Claims 1-4, 11-12, 14-15, and 19-22 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specifications, while being enabling for the septically exemplified stop-codon mutations at position 279 of SEQ ID NO: 2 and positions 233 and 285 of SEQ ID NO: 4, does not reasonably provide enablement for the full scope of the claimed invention.
An “analysis of whether a particular claim is supported by the disclosure in an application requires a determination of whether that disclosure, when filed, contained sufficient information regarding the subject matter of the claims as to enable one skilled in the pertinent art to make and use the claimed invention.” MPEP 2164.01. “A conclusion of lack of enablement means that. . . the specification, at the time the application was filed, would not have taught one skilled in the art how to make and/or use the full scope of the claimed invention [i.e. commensurate scope] without undue experimentation.” In re Wright, 999 F.2d 1557,1562, 27 USPQ2d 1510, 1513 (Fed. Cir. 1993); MPEP 2164.01.
In In re Wands, 858 F.2d 731,8 USPQ2d 1400 (Fed. Cir. 1988), several factors implicated in determination of whether a disclosure satisfies the enablement requirement and whether any necessary experimentation is “undue” are identified. These factors include, but are not limited to:
(A) The breadth of the claims;
(B) The nature of the invention;
(C) The state of the prior art;
(D) The level of one of ordinary skill;
(E) The level of predictability in the art;
(F) The amount of direction provided by the inventor;
(G) The existence of working examples; and
(H) The quantity of experimentation needed to make or use the invention based on the content of the disclosure. In re Wands, 858 F.2d 731,737, 8 USPQ2d 1400, 1404 (Fed. Cir. 1988). No single factor is independently determinative of enablement; rather “[I]t is improper to conclude that a disclosure is not enabling based on an analysis of only one of the above factors while ignoring one or more of the others.” MPEP 2164.01. Likewise, all factors may not be relevant to the enablement analysis of any individual claim.
Claim 1 broadly recites a tobacco plant comprising one or both of: (I) an endogenous gene including a coding region encoding SEQ ID NO: 2 or an amino acid sequence having at least 95% identity with SEQ ID NO: 2; and (ii) an endogenous gene including a coding region encoding SEQ ID NO: 4 or an amino acid sequence having at least 95% identity with SEQ ID NO: 4, wherein at least one of codons corresponding to the respective amino acid sequence is mutated to a stop codon.
The specification enables only a limited number of specific working embodiments. In Example 1, applicant isolated one NtCLCa-S mutant, 220S, in which W279 of SEQ ID NO: 2 was mutated to a stop codon, and two NtCLCa-T mutants, 222T and 223T, in which W233 and W285 of SEQ ID NO: 4 were mutated to stop codons (pa0206-0221). The phenotypic data are based on selected combinations of these particular alleles (pa0222-0239, and Figs 1-4).
The claim scope is broader that the working examples, Frist, claim 1 encompasses amino acid sequences having at least 95% identity with SEQ ID NO: 2 or SEQ ID NO: 4. The specification states that such variants may include deletion, substitutions, insertions, or additions (pa0083-0087). However, the specification does not provide sufficient guidance identifying which amino acid changes within the broad ≥95% identity genus preserve NtCLCa nitrate transport function or still produce the required nitrate-decrease phenotype after introduction of a stop codon (pa 0080-0082, and pa0096).
Second, claim 1 broadly recites that “at least one” codon corresponding to the amino acid sequence is mutated to a stop codon. This encompasses numerous possible stop-codon positions. The working examples are limited to W279STOP in NtCLCa-S and W233STOP/W285STOP in NtCLCa -T (pa0219). Although the specification lists additional possible codon positions and frameshifted embodiments (pa0106-0112), it does not provide working data or predictive guidance showing that stop codons across the full claimed scope would reliably produce the desired nitrate phenotype without undue experimentation.
The unpredictability is confirmed by the specification itself. Applicant states that the lack of nitrite increase was unexpected compare to Arabidopsis CLCa mutant results (pa0234). Applicant also states that increased free amino acid content was unexpected compare to Arabidopsis CLCa mutant results (p0238). These statements show that the relationship between CLCa mutation, nitrate reduction, nitrite level, amino acid level, and plant growth is not fully predictable across the full claimed scope.
Claims 2-4 do not cure the enablement deficiency. Claim 2 narrows the stop codon to positions 233-285, but still covers numerous possible stop-codon positions in that region and ≥95% identity and stop-codon scope unchanged. Claim 4 recites homozygosity, but does not limit the claim to the specific enabled W279STOP, W233STOP, or W285STOP alleles.
Claims 11-12 further require functional properties, including decreased nitrate content, equivalent nitrite content, increased amino acid content, and at least 80% nitrate decrease. The specification provides such data only for selected tested genotypes (pa0231-0239, and figs 1-4). The specification does not enable a person of ordinary skill in the art to predict which untested ≥95% identity variants and stop-codon positions would satisfy these functional properties without screening and phenotypic testing.
Claim 14 limits the tobacco plant to Nicotiana tabacum, but this does not cure the deficiency because the claim still encompasses broad ≥95% identity variants and numerous possible stop-codon positions.
Claim 15 is also not enables. Claim 15 recites a tobacco plant comprising one or both endogenous genes encoding a polypeptide lacking an amino acid sequence form an amino acid residue between positions 233 to 285 to the C-terminal amino acid residue of SEQ ID NO: 2 or 4, or sequence having at least 95% identity thereto. Although the specification provides examples of specific truncations, claim 15 covers a broad genus of truncations beginning anywhere between positions 233 and 285, in either or both genes, including ≥95% identity variants. The specification does not provide sufficient guidance to predict which truncations and variants across this genus would produce the desired nitrate phenotype without undue experimentation.
Claim 19-22 are directed to leaf tobacco, cured leaf, cut filler, powder, sheet, stem, granule, extract, and tobacco products derived from the tobacco plant of claim 1. These claims depend on the non-enabled tobacco plant genus of claim 1 and therefore are not enabled for the same reasons.
Accordingly, considering the breadth of the claims, the limited working examples, the broad ≥95% identity sequence genus, the numerous possible stop-codon/truncation embodiments, and the unpredictability of the nitrate/nitrate/amino-acid phenotype, the specification does not enable the full scope of claims 1-4, 11-12, 14-15, and 19-22 without undue experimentation.
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.
Claims 1-4, 9, 11-14, and 19-22, and 24 are rejected under 35 U.S.C. §103 as being unpatentable over Bovet (Lucien Bovet et. al., US10563215B2, Application: 2013-12-19, Publication: 2015-11-05).
Claim 1 recites a tobacco plant comprising one or both of: (i) an endogenous gene including a coding region encoding SEQ ID NO: 2 or an amino acid sequence having at least 95% identity with SEQ ID NO: 2; and (ii) an endogenous gene including a coding region encoding SEQ ID NO: 4 or an amino acid sequence having at least 95% identity with SEQ ID NO: 4, wherein at least one of codons corresponding to the respective amino acid sequence is mutated to a stop codon. Claim 9 recites the tobacco plant according to claim 1, wherein in the nucleic acid included in the endogenous gene of (i), an amino acid corresponding to position 163 of SEQ ID NO: 2 encoded by the nucleic acid is glycine. Claim 13 recites the tobacco plant according to claim 1, wherein the tobacco plant is a mutant or a gene-modified variant. Claim 14 recites the tobacco plant according to claim 1, wherein the tobacco plant is Nicotiana tabacum.
Bovet teaches tobacco plants that CLC genes exist in Nicotiana tabacum, and further teaches reducing expression of these genes in tobacco plants to reduce nitrate levels (p2, line52-65). Bovet teaches modulating the expression of activity of tobacco CLC polypeptides, including tobacco CLC sequences corresponding to the SEQ ID NO: 5 (CLCNt2-s) and SEQ ID NO: 6 (CLCNt2-t)(Abstract, pa131-134, Table 1 and 2). Bovet further teaches SEQ ID NO: 5 and SEQ ID NO: 6, are identical to the instant claimed SEQ ID NO: 2 and SEQ ID NO: 4 respectively (pa131-134) (see alignment below). Bovet further teaches that expression of polypeptide may be modulated by non-transgenic means, such as creating a mutation in a gene, including chemical mutagenesis, EMS mutagenesis, radiation mutagenesis, and targeted mutation methods (pa33, line 25-35). Bovet also teaches that screening may be performed to identify mutations that create premature stop codons or otherwise non-functional genes (pa33, line 36-59; pa38, line 36-49).
Bovet does not expressly exemplify the presently claimed tobacco plant having a stop codon in the endogenous gene encoding SEQ ID NO: 2 or SEQ ID NO: 4. However, Bovet teaches the same tobacco CLCNt2 target sequence and teaches reducing or modulating their expression/function to reduce nitrate/TSNA. Bovet also expressly teaches creating mutations, including premature stop codons, to produce non-functional genes.
Therefore, it would have been obvious to one of ordinary skill in the art to introduce or select a premature stop-codon mutation in the endogenous tobacco CLC gene taught by Bovet in order to reduce or eliminate CLC function and thereby reduce nitrate accumulation in tobacco leaves.
Accordingly, the claimed invention in claims 1, 9, and 13-14 as a whole is a prima facies obvious over Bovet.
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Claim 2 further recites that the sop codon is located at a codon corresponding to amino acids 233 to 285 of SEQ ID NO: 2 and/or SEQ ID NO: 4. Claim 3 recites the tobacco plant according to claim 1, comprising both endogenous genes of (i) and (ii). Claim 4 recites the tobacco plant according to claim 1, wherein the endogenous gene of (i) and/or the endogenous gene of (ii) is a homozygote.
For the same reasons set forth above with respect to claim 1, Bovet teaches the same tobacco CLC target sequences and teaches creating mutations, including premature stop codons, to obtain non-functional genes. selecting a stop-codon mutation within the coding region of the known CLC gene would have been a routine way to disrupt CLC formation. A premature stop codon within the recited region would predictably truncated the CLC protein, and reduce or eliminate its activity.
Bovet teaches homozygous CLCNt2-s G163R mutant plants and homozygous CLCNt2-s G163R mutant plants in the description of FIGs. 5 and 6 (pa7, line 37-58). Bovet therefore teaches selecting homozygous tobacco plant carrying mutations in the relevant tobacco CLC genes.
Accordingly, the claimed invention in claims 2-4 as a whole is a prima facies obvious over Bovet.
Claim 11 recites that the tobacco plant of claim 1 has one or more properties selected from decreased nitrate content, equivalent nitrite content, and increased amino acid content compared to a control.
Bovet teaches tobacco plants in which nitrate content is decreased compare to control tobacco plants by modulating tobacco CLC gene expression/function (Claim 1). Accordingly, claim 11 is obvious over Bovet.
Claim 12 recites the tobacco plant according to claim 11, wherein a nitrate content is decreased by at least 80% compared to the control.
Bovet teaches tobacco plants having substantially reduced nitrate content as compared to wild-type control plants. As shown in FIG 2A (also see below), the CLCNt2-RNAi plants exhibit nitrate levels of approximately 10-15% of wild-type plants, corresponding to a nitrate reduction greater than 80%. Although Bovet achieves this reduction through RNAi-mediated suppression of CLC gene expression rather than through stop-codon mutations or truncated proteins, Bovet nevertheless teaches that reduction of CLC gene function/activity results in markedly decreased nitrate accumulation. It would have been obvious to a person of ordinary skill in the art that alternative loss-of-function approaches, including introduction of stop codons or truncating mutations in the endogenous CLC genes as recited claim 1 would likewise reduce CLC function and produce substantially reduced nitrate content. Accordingly, claim 12 is obvious over Bovet.
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Claim 15 recites a tobacco plant comprising one or both endogenous gene including, as a coding region, a nucleotide sequence encoding a polypeptide lacking an amino acid sequence from an amino acid residue between positions 233 to 285 to a C-terminal amino acid residue of SEQ ID NO: 2 or SEQ ID NO: 4, or sequences having at least 95% identity thereto.
For the same reasons set forth above with respect to claims 1 and 2, Bovet teaches the same tobacco CLC sequences and teaches mutations that create premature stop codons or otherwise non-functional genes. As discussed above for claim 2, introducing a premature stop codon within the region corresponding to amino acids 233 to 285 would predictably produce a truncated CLC polypeptide lacking the downstream C-terminal amino acid sequence, as recited in claim 15.
Accordingly, claim 15 is obvious over Bovet.
Claim 19 recites to leaf tobacco harvested from the tobacco plant according to claim 1. Claim 20 recites to a cured leaf generated from the leaf tobacco according to claim 19. Claim 21 recites to a cut filler, powder, sheet, stem, granule, or extract produced from the cured leaf according to claim 20. Claim 22 recites to a tobacco product comprising the cured leaf according to claim 20. Claim 24 recites to a tobacco product comprising the cut filler, powder, sheet, stem, granule, or extract according to claim 21.
Claims 19-22 and 24 are reject for the same reasons set forth above with respect to claim 1. Bovet further teaches harvesting plant material, such as one or more leaves, from the mutant, non-naturally occurring, or transgenic tobacco plant, curing the harvested plant material for a period of time, and identifying cured plant material having reduced nitrate content compared to a control plant (pa46, lin16-30). Bovet therefore teaches or suggests leaf tobacco harvested from the claimed tobacco plant, cured leaf generated from the leaf tobacco, tobacco materials derived from the cured leaf, and tobacco products comprising the cured leaf or tobacco materials. Accordingly, claims 19-22 and 24 would have been obvious.
Conclusion
No claims are allowed.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to YANXIN SHEN whose telephone number is (571)272-7538. The examiner can normally be reached Monday-Friday.
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If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Amjad A Abraham can be reached at (571)272-7058. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/YANXIN SHEN/ Examiner, Art Unit 1663
/WEIHUA FAN/ Primary Examiner, Art Unit 1663
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