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 .
Claims 1-4, 6-7, 9, 11, 13, 15, 17, 19, 22-25, 35, 38-41, and 43-60 are pending and examined on their merit herein.
Claim Rejections - 35 USC § 112
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 54-56 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 54 is indefinite since it is dependent on canceled claims. Dependent claims 55 and 56 are included in this rejection.
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.
Claims 1-4, 6-7, 9, 11, 13, 15, 17, 19, 22-25, 35, 38-41, and 43-60 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification, while being enabling for recombinant DNA molecule encoding a protein comprising SEQ ID NO: 18, 38, or 42, does not reasonably provide enablement for any of the mutation combinations in the proteins having 70% identity with any of SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, or 16, and so on. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and/or use the invention commensurate in scope with these claims.
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.
The claims are broad in encompassing nucleic acid sequence encoding a protein having at least 70% sequence identity to SEQ ID NOs: 2, 4, 6, 8, 10, 12, 14, and 16, and the protein comprises at least one amino acid substitution as compared to SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, or 16, mutations selected from 141Y/F/H, etc (SEQ ID NO: 2 numbering), and wherein the protein is required to have triketone dioxygenase (TDO) activity.
The claims are broad in scope at least for the following reasons: 1) a protein having 70% identity with SEQ ID NO: 2—which has 351 amino acids—would have as many as 105 amino acid substitutions, insertions, deletions, or additions, anywhere in the protein in any combinations. 2) furthermore, since SEQ ID NO: 4, 6, 8, 10, 12, 14, or 16, each has ~62-68% identity with SEQ ID NO: 2, a protein having 70% identity with any of SEQ ID NO: 4, 6, 8, 10, 12, 14, or 16 would potentially have as low as 43% identity with SEQ ID NO: 2. Thus, the genus of proteins and their variants encompassed by the claims is broad and has a diverse range of structures.
In contrast to the broad scope, the Specification has taught the rice HPPD Inhibitor Sensitive 1 (HIS1) gene which has been previously found to confer tolerance to triketone herbicides, including mesotrione (Example 1); and that rice HIS1 gene encodes the protein of amino acid sequence SEQ ID NO: 2. The Specification has taught ectopic overexpression of rice HIS1 (with optimized codon for dicot) in soybean conferring field tolerance to mesotrione of up to 16x of field rate (Example 1).
The Specification has taught homolog genes of rice HIS1 from corn, sorghum, wheat, and Setatria italica (Foxtail millet), etc., that encodes the amino acid sequences SEQ ID NO: 4, 6, 8, 10, 12, 14, or 16 (Table 1). As mentioned above, these homolog proteins share about 62 to 68% identity to SEQ ID NO: 2 (Table 1).
The Specification has taught ectopic expression of the homologs also in soybean. The Specification has taught that “while homologs from com, sorghum, showed reduced but still significant injury, those from wheat, Andropogon gerardii, and Coix lacryma-jobi did not seem to provide any protection against mesotrione.” (Example 2, and p. 37, [0112]) Note that these homologous proteins are cited in claim 1, for example, wheat, which is SEQ ID NO: 10).
Regarding the mutations, the Specification has taught variants of the rice HIS1 (TDO) protein, e.g., variant “D08”, which has 20 amino acid substitutions compared with SEQ ID NO: 2 (as set forth in SEQ ID NO: 18), or “B09”, as set forth in SEQ ID NO: 26, having 22 amino acid substitutions relative to SEQ ID NO: 2. (Example 4, Table 3) The Specification has taught that while the five variants, SEQ ID NO: 18, 20, 22, 24, 26, showed increased heat stability, however, “all five also showed reduced enzyme activity in vitro”. ([0118]). Particularly, SEQ ID NO: 26 only has 0.2% of the wild type HIS1 activity (or a 418-fold decrease in enzymatic activity; see data in Table 3).
The Specification has taught a further mutational optimization of the most promising variant “D08”, e.g., with variants presented in Table 4, Example 4, and SEQ ID NO:18 (which is D08), 28, 30, 32, 34, 36, 38, 40, 42, or 44 (cited in claim 38). Transgenic cotton plants showed that two of the variants, D08 and TDO_6, showed improved herbicide tolerance at the higher temperature, suggesting the enhanced heat stability of the enzyme translates to improved herbicide tolerance in cotton. (Example 4). These two variants have the sequences of SEQ ID NO: 18 and 38, respectively.
The Specification has taught among the homologs tested, only the maize TDO (SEQ ID NO: 4) showed limited activity in providing tolerance to triketone herbicides (Example 5). Baed on the best understanding from the multiple sequence alignment, and the mutational studies discussed above, the Specification has taught a hypothesis regarding the structural determinant of the herbicide-metabolizing activity. “To test the hypothesis, the four amino acids of sorghum TDO corresponding to amino acid positions 230, 301, 332, and 335 in rice TDO were replaced with those of rice TDO”. (Example 5). The so-called SwM variant comprises the combination of substitutions P230T-A301S-L332R-V335I, in which the wild-type sorghum residues at positions 230, 310, 332, and 335 were replaced with those from rice TDO. However, the result showed that the modified sorghum TDO variant designated as SwM had about 17% activity compared to the rice TDO protein. (Example 5, [0132]).
Lastly, the Specification postulated “surveying other TDOs based on their similarity to the active site of rice TDO, rather than the similarity of the entire protein, may provide a valid strategy to identify naturally active or other variants.” ([0136], the last paragraph of the Specification).
In summary, the Specification has taught that the natural form of rice TDO (HIS1, SEQ ID NO: 2) provides field tolerance to triketon herbicides in transgenic soybean; and two variants with improved heat stability and herbicide tolerance.
However, the Specification has not provided enabling teachings of how to make and/or use the:
1) proteins or protein variants with any one of the substitutions such as a 141Y/F/H, etc., in any of SEQ ID NO:2, 4, 6, 8, 10, 12, 14, or 16, or proteins having 70% identity; or further comprising any of mutations such as L11;
2) proteins or protein variants with any of 230T, 301S, 332R, or 335I in any of SEQ ID NO: 4, 6, 8, 10, 12, 14, or 16, or proteins having 70% identity;
3) proteins or protein variants with 230T-301S-332R-335I in any of SEQ ID NO: 4, 6, 8, 10, 12, 14, or 16, or proteins having 70% identity;
4) proteins or protein variants with 230T-301S-332R-335I in any of SEQ ID NO: 4, 6, 8, 10, 12, 14, or 16, or proteins having 70% identity;
5) the protein with 50% identity to SEQ ID NO: 2 and comprises any one or more substitutions I11L, A18K, K32Q, S35P, L48I, S54G, V82I, S89D, N116H, Q120E, I133V, N167H, T173C, L199F, A204T, T230G, S242G, E256P, C274S, R279K; or the combinations; or
6) the protein as set forth in SEQ ID NO: 48;
With regard to any specific tolerance to herbicides.
The Specification, and the state of the art at the time of filing, has not provided enabling guidance for the structural determinant of the specific levels of tolerance toward the specific array of herbicides; or how the amino acid variations would impact the enzymatic activity, specificity, as well as heat stability. The Specification has not provided enabling guidance on how to make and combine as many as 105 amino acid substitutions, insertions, deletions, or additions, anywhere in the proteins to have the required herbicide tolerance level, specificity, and stability.
Firstly, as the Specification shown, even when proteins from other plant species, even monocot species, are identified as triketone dioxygenase, they do not confer tolerance to mesotrione ([0112]).
Secondly, the Specification has hypothesized, that since rice TDO (HIS1, SEQ ID NO: 2) is uniquely having the mesotrione tolerance activity, engineering the homolog proteins, such as the sorghum protein SEQ ID NO: 6, to have the same amino acids as SEQ ID NO: 2 at the critical positions, e.g., 230, 301, 332, and 335, would turn the non-active sorghum protein into a function TDO. However, the attempt was not successful: the Specification showed that modified sorghum protein “SwM variant” with the combination of substitutions P230T-A301S-L332R-V335I, had about 17% activity compared to the rice TDO protein. (Example 5, [0132]). This was later confirmed in 2023, when Duff (Biochimica et Biophysica Acta (BBA)-General Subjects 1868.2 (2024): 130504) taught:
“When these four rice TDO amino acids, T230, S301, R332 and I335 are introduced at the proper locations in sorghum TDO (which is not active to mesotrione), a small level of mesotrione activity was introduced to the protein, suggesting they are critical for activity and providing an opening for engineering additional TDO proteins with herbicide resistance. For example, since corn TDO is the only other protein we tested that has activity, it would be of interest to replace the first three of these amino acids (T230, S301, and R332) into maize TDO. However, since the activity of the mutant sorghum TDO was lower than either the native maize or rice TDO enzymes, it likely would not protect maize plants more efficiently from mesotrione and further modification of the enzyme will be necessary to engineer TDO enzymes with greater activity.”
And Duff in Fig. 6:
PNG
media_image1.png
520
514
media_image1.png
Greyscale
Showing the specific TDO activity compared with that of rice TDO.
Duff went on, even as of 2023, to admit that “the uniqueness of rice TDO suggests that surveying other TDOs based on their similarity to the active site of rice TDO, rather than the similarity of the entire protein, would be a valid strategy to identify natural active variants.”
These evidences prove, that Specification, and the state of the art at the time of filing, only began to work toward understanding the structural determinant for the mesotrione tolerance activity which is unique to rice TDO. The levels of predictability, between any specific amino acid substitutions or combinations, with the specific mesotrione metabolizing activity, was low at the time of the filing of the instant application.
The lack of sufficient understanding regarding the structural determinant is further demonstrated in other teachings. For example, Takamura (Plant physiology 187.2 (2021): 816-828) taught that while HSL genes (e.g., those represented by the instant SEQ ID NO: 4, 6, 8, etc.,) are highly conserved among poaceous species, suggestive of common and important roles of HSL gene products in major crops, the natural substrate (or substrates) of HIS1 and HSL proteins remains poorly understood. Furthermore, Takamura taught that while TE (trinexapac-ethyl) is a general inhibitor of 2-oxoglutarate (2OG)/Fe(II)-dependent dioxygenase, it was unexpectedly found that TE is recognized as a substrate by HIS1 (p. 818). Takamura further taught that none of the single mutants examined (F140H, L204F, and F298L) manifested TE metabolizing activity. Both double mutants containing F140H did acquire the ability to metabolize TE, and the triple mutant showed a level of activity similar to that of HIS1. Takamura taught that the effect of the F298L mutation was unexpected because OsHSL2 also has Phe at the corresponding position. Takamura suggested that minor structural changes induced by amino acid substitutions in the putative substrate pocket of OsHSL1 lead to marked changes in protein function, which has not been adequately understood or illustrated at the filing of the instant application.
Since the Specification has not provided adequate guidance as to which one those substitutions should or should not be combined with other mutations, a skilled artisan would not be able to readily appreciate if the combination would have desired increase in enzymatic activity, specificity, or protein stability.
In the absence of guidance from either the instant disclosure or the art, it would require trial and error experimentation for a skilled artisan to identify the combinatorial mutants with increased activity.
Thus, in view of the unpredictability associated with combinatorial substitutions in a protein, the lack of enabling guidance from either the instant disclosure or the art, and breath and diversity of the embodiments encompassed by the claimed genus, the lack of sufficient working examples, and the level of the art at the time of the invention, one of ordinary skill in the art must rely on undue trial and error experimentation to make and test the numerous polypeptides having multiple amino acid substitutions, in order to make and/or use the invention within the full scope of these Claims.
For at least this reason, the Specification does not teach a person with skill in the art how to make and/or use the subject matter within the full scope of these Claims.
Claim Rejections - 35 USC § 102
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 (i.e., changing from AIA to pre-AIA ) 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 the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 1, 2, 4, 23-24, 40-41, and 43-46 are rejected under 35 U.S.C. 102(a)(1) and (a)(2) as being anticipated by Coffin (US PGPUB US 20160244777 A1, published 2016).
Claim 1 is drawn to a recombinant DNA molecule comprising a nucleic acid sequence encoding a protein having triketone dioxygenase (TDO) activity, wherein the protein has at least 70% sequence identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 2, 4, 6, 8, 10, 12, 14, and 16, wherein the protein comprises at least one amino acid substitution as compared to SEQ ID NO: 2, 4, 6, 8, 10, 12, 14, or 16, and wherein the protein comprises:
a Y, F, or H at the position corresponding to position 141 of SEQ ID NO: 2;
an A, I, L, S, T, V or G, at the position corresponding to position 229 of SEQ ID NO: 2;
a T, A, L, V, I, S, or G, at the position corresponding to position 230 of SEQ ID NO: 2;
an A, I, L, or V, at the position corresponding to position 299 of SEQ ID NO: 2;
an S at the position corresponding to position 301 of SEQ ID NO: 2;
an R at the position corresponding to position 332 of SEQ ID NO: 2;
an I at the position corresponding to position 335 of SEQ ID NO: 2;
or
a combination of any thereof;
provided that:
(i) when a T, A, L, V, I or S is present at the position corresponding to position 230, a G is present at the position corresponding to position 229 of SEQ ID NO: 2; and
(ii) when an A, L, V, I, T, or S is present at the position corresponding to 229, a G is present at the position corresponding to position 230 of SEQ ID NO: 2.
Note that the claim specifies amino acid substitutions at positions corresponding to positions 141, 229, 230, 299, 301, 332 and 335 as the alternatives (“or”), and any combinations; and one of the amino acids corresponding to positions 229 and 230 has to be a G when the other is a T, A, L, V, I or S.
Regarding claim 1, Coffin discloses a recombinant DNA construct comprising a promoter that is functional in said plant cell and that is operably linked to a protein coding DNA encoding a protein having an amino acid sequence selected from the group consisting of SEQ ID NO: 760 through SEQ ID NO: 67778. (Claims 1, 2 and 3). It is noted that the prior art SEQ ID NO: 4755 is a protein sequence having 91% identity with the instant SEQ ID NO: 2; and having an H at position 140 which is corresponding to the position 141 or the instant SEQ ID NO: 2; and an S at the position corresponding to 230 of instant SEQ ID NO: 2 while a G at 229; an L at 299, S at 301, R and I at positions corresponding to positions 332 and 335 of the instant SEQ ID NO: 2. See sequence alignment below.
In summary, the prior art recombinant DNA comprises a DNA sequence encoding a protein, 1) having more than 70% identity with the instant SEQ ID NO: 2, and 2) at the corresponding positions with SEQ ID NO: 2, having 141H, G229/S230, L299, S301, R332 and I 335. Therefore, Coffin discloses at least one of the embodiments claimed in Claim 1.
Sequence alignment of the instant SEQ ID NO: 2 (query) with Coffin SEQ ID NO: 4755. Note the relevant positions are shown in bold font and underlined.
RESULT 3
US-14-757-273-4755
(NOTE: this sequence has 2 duplicates in the database searched.
See complete list at the end of this report)
Sequence 4755, US/14757273
Patent No. 10167482
GENERAL INFORMATION
APPLICANT: Coffin, Marie
TITLE OF INVENTION: GENES AND USES FOR PLANT ENHANCEMENT
FILE REFERENCE: 38-21(54866)B
CURRENT APPLICATION NUMBER: US/14/757,273
CURRENT FILING DATE: 2015-12-10
PRIOR APPLICATION NUMBER: 12/157,153
PRIOR FILING DATE: 2008-06-05
PRIOR APPLICATION NUMBER: 60/933,428
PRIOR FILING DATE: 2007-06-06
NUMBER OF SEQ ID NOS: 67778
SEQ ID NO 4755
LENGTH: 350
TYPE: PRT
ORGANISM: Oryza sativa
Query Match 92.3%; Score 1679.5; Length 350;
Best Local Similarity 91.2%;
Matches 320; Conservative 17; Mismatches 13; Indels 1; Gaps 1;
Qy 1 MADESWRAPAIVQELAAAGVEEPPSRYLLREKDRSDVKLVAAELPEPLPVVDLSRLDGAE 60
||||||| |||||||||||||||||||:| |||||| :|||||||||:|||||||| ||:
Db 1 MADESWRTPAIVQELAAAGVEEPPSRYVLGEKDRSD-ELVAAELPEPIPVVDLSRLAGAD 59
Qy 61 EATKLRVALQNWGFFLLTNHGVEASLMDSVMNLSREFFNQPIERKQKFSNLIDGKNFQIQ 120
|| ||| |||||||||||||||| |||| |:||:|||||||||||:||||||||||||::
Db 60 EAAKLRAALQNWGFFLLTNHGVETSLMDDVLNLAREFFNQPIERKRKFSNLIDGKNFQVE 119
Qy 121 GYGTDRVVTQDQILDWSDRLHLRVEPKEEQDLAFWPDHPESFRDVLNKYASGTKRIRDDI 180
|||||||||||||||||||| ||||||||::||||||||||||||||:||| ||||||||
Db 120 GYGTDRVVTQDQILDWSDRLFLRVEPKEERNLAFWPDHPESFRDVLNEYASRTKRIRDDI 179
Qy 181 IQAMAKLLELDEDYFLDRLNEAPAFARFNYYPPCPRPDLVFGIRPHSDGTLLTILLVDKD 240
:|||:||| |||||| ||||:||| |||||||||||||||||:||||||:| ||||||||
Db 180 VQAMSKLLGLDEDYFFDRLNKAPALARFNYYPPCPRPDLVFGVRPHSDGSLFTILLVDKD 239
Qy 241 VSGLQVQRDGKWSNVEATPHTLLINLGDTMEVMCNGIFRSPVHRVVTNAEKERISLAMLY 300
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 240 VSGLQVQRDGKWSNVEATPHTLLINLGDTMEVMCNGIFRSPVHRVVTNAEKERISLAMLY 299
Qy 301 SVNDEKDIEPAAGLLDENRPARYRKVSVEEFRAGIFGKFSRGERYIDSLRI 351
|||||||||||||||||||||||||||||||||||||||||||||||||||
Db 300 SVNDEKDIEPAAGLLDENRPARYRKVSVEEFRAGIFGKFSRGERYIDSLRI 350
Claim 2 is drawn to the recombinant DNA, wherein the protein 230T, 301S, 332R, 335I; or a combination of any thereof. Since the amino acids are specified as alternatives, Coffin at least discloses the 301S-332R-335I combination (see above).
Regarding claim 4, Coffin discloses the embodiment of H141F, at least. (See above).
Regarding claims 23-24, Coffin discloses the protein having an I at position 48, for example (See above).
Regarding claims 40-41, Coffin discloses the DNA molecule comprising a heterologous promoter functional in plant cells (see above, Coffin at Claim 1, e.g.).
Regarding claims 43-46, Coffin discloses the stable incorporation of the recombinant DNA into a plant cell genome (Claim 1), the encoded protein, as well the DNA construct, transgenic plant.
Therefore, claims 1, 2, 4, 23-24, 40-41, and 43-46 are anticipated by Coffin.
Claims 1-2, 40-41, 43-48 and 53-60 are rejected under 35 U.S.C. 102(a)(1) and (a)(2) as being anticipated by TOZAWA (US PGPUB US 20200048315 A1, published 2/13/2020, priority filing date 2/9/2018).
Regarding claim 1, TOZAWA discloses a rice HIS1-like gene, encoding a HSL1 protein the amino acid sequence of SEQ ID NO: 4, which is 87% identical with the instant SEQ ID NO: 2 (see below). Note that position 140 of rice HSL1 protein corresponds to position 141 of rice HIS1 (instant and TOZAWA SEQ ID NO: 2) (see alignment and also TOZAWA Table 7).
TOZAWA discloses, to increased resistance to a 4- HPPD inhibitor herbicide, mutating, in an HSL protein of a plant cell, position 140 of an amino acid sequence of SEQ ID NO : 4 or an amino acid corresponding to the position to a basic amino acid; and regenerating a plant from the plant cell in which amino acid mutation is introduced; wherein the basic amino acid is histidine, lysine, or arginine (Claims 2-3). In addition, TOZAWA discloses a single-site mutant of F140H was expressed in Arabidopsis thaliana, and conferring resistance against sulcotrione, mesotrione, or tembotrione ( [0245, Example 5), introduced by transformation with a recombinant DNA construct (Example 4). Therefore, TOZAWA discloses recombinant DNA encoding a protein having more than 70% identity with the instant SEQ ID NO: 2 and a substitution (compared with the corresponding wildtype sequence) at a position corresponding to 141 od the instant SEQ ID NO: 2, wherein the amino acid is an H.
Therefore, claim 1 is anticipated by TOZAWA.
Regarding claim 2, TOZAWA discloses mutating rice HSL1 with a S229T substitution (Example 3, [0161], for example); which corresponding to the instant 230T. Note that at the position corresponding to the instant 229, which is position 228 of rice HSL1 (TOZAWA SEQ ID NO: 1) the native amino acid is a G. Therefore, TOZAWA discloses the 230T mutation which also satisfies the limitation of the instant claim 1 “(i) when a T, A, L, V, I or S is present at the position corresponding to position 230, a G is present at the position corresponding to position 229 of SEQ ID NO: 2”.
Regarding claims 40-48, TOZAWA discloses the recombinant vector, comprising the mutated HSL1 gene linked to a plant promoter (35S promoter), transgenic plant (Arabidopsis, Example 5; rice, Example 6), wherein the transformed plants are resistant to HPPD-inhibitor herbicides, e.g., mesotrione (Example 5). Regarding claims 53-60, TOZAWA discloses the corresponding method of making the transgenic plants, and the selection process of obtaining the resistant plant, as discussed above (Examples 5 and 6, e.g.)
Therefore, claims 1-2, 40-41, 43-48 and 53-60 are anticipated by TOZAWA.
Alignment of the instant SEQ ID NO: 2 (Query) with TOZAWA SEQ ID NO: 4 (Sbjct):
Score:634 bits(1635), Expect:0.0,
Method:Compositional matrix adjust.,
Identities:305/351(87%), Positives:329/351(93%), Gaps:1/351(0%)
Query 1 MADESWRAPAIVQELAAAGVEEPPSRYLLREKDRSDVKLVAAELPEPLPVVDLSRLDGAE 60
MADESWR PAIVQELAAAGVEEPPSRY+L EKDRSD +LVAAELPEP+PVVDLSRL GA+
Sbjct 1 MADESWRTPAIVQELAAAGVEEPPSRYVLGEKDRSD-ELVAAELPEPIPVVDLSRLAGAD 59
Query 61 EATKLRVALQNWGFFLLTNHGVEASLMDSVMNLSREFFNQPIERKQKFSNLIDGKNFQIQ 120
EA KLR ALQNWGFFLLTNHGVE SLMD V+NL+REFFNQPIERK+KFSNLIDGKNFQ++
Sbjct 60 EAAKLRAALQNWGFFLLTNHGVETSLMDDVLNLAREFFNQPIERKRKFSNLIDGKNFQVE 119
Query 121 GYGTDRVVTQDQILDWSDRLHLRVEPKEEQDLAFWPDHPESFRDVLNKYASGTKRIRDDI 180
GYGTDRVVTQDQILDWSDRL LRVEPKEE++LAFWPDHPESFRDVLN+YAS TKRIRDDI
Sbjct 120 GYGTDRVVTQDQILDWSDRLFLRVEPKEERNLAFWPDHPESFRDVLNEYASRTKRIRDDI 179
Query 181 IQAMAKLLELDEDYFLDRLNEAPAFARFNYYPPCPRPDLVFGIRPHSDGTLLTILLVDKD 240
+QAM+KLL LDEDYF DRLN+APA ARFNYYPPCPRPDLVFG+RPHSDG+L TILLVD+D
Sbjct 180 VQAMSKLLGLDEDYFFDRLNKAPALARFNYYPPCPRPDLVFGVRPHSDGSLFTILLVDED 239
Query 241 VSGLQVQRDGKWSNVEATPHTLLINLGDTMEVMCNGIFRSPVHRVVTNAEKERISLAMLY 300
V GLQ+QRDGKW NV+ TP+TLLINLGDTMEV+CNGIFRSPVHRVVTNAE+ERISLAM Y
Sbjct 240 VGGLQIQRDGKWYNVQVTPNTLLINLGDTMEVLCNGIFRSPVHRVVTNAERERISLAMFY 299
Query 301 SVNDEKDIEPAAGLLDENRPARYRKVSVEEFRAGIFGKFSRGERYIDSLRI 351
SVNDEKDI PAAGLLDENRPARYRKVSV EFRAGI GKFSR ERYIDSL+I
Sbjct 300 SVNDEKDIGPAAGLLDENRPARYRKVSVGEFRAGIIGKFSRRERYIDSLKI 350
Conclusion
No claims are allowed.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to WEIHUA FAN whose telephone number is (571)270-0398. The examiner can normally be reached Monday-Friday, 9-5.
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WEIHUA . FAN
Primary Examiner
Art Unit 1663
/WEIHUA FAN/Primary Examiner, Art Unit 1663