Plants with Modified Deoxyhypusine Synthase Genes

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 without traverse of Group II in the reply filed on 7/23/2026 is acknowledged. The election is made FINAL. Applicant is reminded that upon the cancelation of claims to a non-elected invention, the inventorship must be corrected in compliance with 37 CFR 1.48(a) if one or more of the currently named inventors is no longer an inventor of at least one claim remaining in the application. A request to correct inventorship under 37 CFR 1.48(a) must be accompanied by an application data sheet in accordance with 37 CFR 1.76 that identifies each inventor by his or her legal name and by the processing fee required under 37 CFR 1.17(i). Status of Claims Claims 11-25 are under examination on the merits. Nucleotide and/or Amino Acid Sequence Disclosures This application contains sequence disclosures that are encompassed by the definitions for nucleotide and/or amino acid sequences set forth in 37 CFR 1.831(b). However, this application fails to comply with the requirements of 37 CFR 1.831-1.834. Sequence identifiers are missing from the amino acid sequences presented in Figure 1A-1I and the amino acid and polynucleotide sequences in figures 7-11. Where a sequence that is set forth in the "Sequence Listing XML" is presented in a drawing, reference must be made to the sequence by use of the sequence identifier (§ 1.832(a)), either in the drawing or in the Brief Description of the Drawings, where the correlation between multiple sequences in the drawing and their sequence identifiers (§ 1.832(a)) in the Brief Description is clear. See MPEP 2412.04. If the sequences in figures 1, 7, 8, 9, 10 & 11 are not present in the sequence listing file, they must be provided. SEQ ID NO: 159 in figure 11 is appropriately referenced, but all other polynucleotide sequences longer than 10 bases and amino acid sequences longer than 4 amino acids should also be presented with their sequence ID numbers. The brief description of Figure 3 describes the HELPTE domain and the amino acid sequence HEPE. If HELPTE is a domain name and not an amino acid sequence it might not require a SEQ ID number; however, HEPE is a sequence of 4 specific amino acid residues. Sequences encompassed by 37 CFR 1.831(b) must be listed in the sequence listing file and referenced by use of the sequence identifier, preceded by "SEQ ID NO:" or the like in the text of the description or claims, even if the sequence is also embedded in the text of the description or claims of the patent application. Full compliance with the sequence rules is required in response to this Office action. A complete response to this Office action must include both compliance with the sequence rules and a response to the issues set forth herein. Failure to fully comply with both of these requirements in the time period set forth in this Office action will be held to be non-responsive. Specification The disclosure is objected to because of the following informalities: The brief description of figure 4 references pink lines and red lines (page 11, lines 7-11). The brief description of figure 5 references yellow lines (page 11, lines 16-19). Since there are no color drawings and no petition for color drawings, reference to colored elements in the provided drawings should be removed. Appropriate correction is required. 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. Claims 11-25 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. All dependent claims are included in the rejection. Claim 11 recites “the” activity of DHS in line 8. There is insufficient antecedent basis for this limitation in the claim, because DHS is highly promiscuous and can catalyze various reactions (see Kaltenegger et al (2021) FEBS Open Bio. 11: 10-25 (Published 11/28/2020, hereafter Kaltenegger); abstract). While the main activity of DHS is the activation of eIF5A, plant DHS are known to be promiscuous with regard to the utilization of aminobutyl donors and acceptors, such as transferring the aminobutyl moiety from spermidine to putrescine to yield homospermidine, an important precursor of pyrrolizidine alkaloids (page 11, left column, paragraph 3-right column, paragraph 1). Because wildtype DHS has multiple activities, it is indefinite which activity is “the” activity that is decreased in the method of claim 11, or if all activity of DHS is required to decrease. Scope of Enablement 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 11-24 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 the method of producing a tomato plant with delayed senescence comprising inducing at least one nucleotide deletion or insertion into at least one copy of a gene encoding deoxyhypusine synthase in a codon for at least one amino acid of a six-amino acid hypervariable region, does not reasonably provide enablement for any insertion or deletion into any DHS gene in any plant wherein the deletion or insertion decreases the activity of DHS so as to delay senescence. 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 the invention commensurate in scope with these claims. Claims 11-16 & 19-24 are drawn to methods of producing a plant with delayed senescence comprising inducing at least one nucleotide deletion or insertion into at least one copy of a gene encoding DHS in a plant in a codon for an amino acid of a six-amino acid hypervariable region, wherein the deletion or insertion decreases the activity of DHS encoded by the gene so as to delay senescence. Claims 17-18 are drawn to a plant or progeny produced by the method. Claims 14, 15 & 16 further require that the senescence is age-related, environmental stress-induced or pathogen-induced senescence respectively. Claims 19, 24 & 25 limit the species of plant. Thus, the claims require a plant comprising 1. at least one nucleotide deletion or insertion into at least one copy of a gene encoding DHS in the recited region, 2. a decreased activity of DHS due to the deletion or insertion, and 3. delayed senescence. The instant specification teaches 35 DHS genes in 34 species, although this number includes a human DHS homolog (table 1). Some species, such as Solanum lycopersicum, may have more than one DHS gene (page 16, lines 25-28). The DHS genes taught by the instant specification do not all have the same number of amino acids in the six-amino acid hypervariable region; see figure 1c or the alignment of instant SEQ ID NO: 39, from Cucurbita pepo, to instant SEQ ID NO: 12, from Brassica napus, below. Sequence 12, PC/TUS2369698A GENERAL INFORMATION APPLICANT: AGRIBODY TECHNOLOGIES, INC. (en) TITLE OF INVENTION: PLANTS WITH MODIFIED DEOXYHYPUSINE SYNTHASE GENES (en) FILE REFERENCE: 117188-5013-WO CURRENT APPLICATION NUMBER: PCT/US23,69698 CURRENT FILING DATE: 2023-07-06 NUMBER OF SEQ ID NOS: 156 SEQ ID NO 12 LENGTH: 368 TYPE: PRT FEATURE: NAME/KEY: source LOCATION: 1..368 QUALIFIERS: mol_type = protein organism = Brassica napus Query Match 84.7%; Score 1654.5; Length 368; Best Local Similarity 84.5%; Matches 306; Conservative 33; Mismatches 22; Indels 1; Gaps 1; Qy 1 MENDNVLSSVHTTVFMKSENLEGKSIKIEGYDFNHGVNYSKLLSSMVSTGFQASNLGDAI 60 || | ||||||:||| :||:|||| |||||||| |||| ||| ||::|||||||||:|| Db 1 MEEDRVLSSVHSTVFKESESLEGKCDKIEGYDFNQGVNYPKLLRSMLTTGFQASNLGEAI 60 Qy 61 EIINQMIDWRLADENIT-EDCSEEEKDPIFRQSTRCKVFLGFTSNLISSGVRDTIRYLVE 119 :|:|||::|||:|| | |||||||||| :|:| :||:||||||||:|||||:||||||: Db 61 DIVNQMLEWRLSDETIAPEDCSEEEKDPAYRESVKCKIFLGFTSNLVSSGVRETIRYLVQ 120 Qy 120 HRMVDVIVTTTGGIEEDLIKCLAPTYKGDFSLPGAVLRSKGLNRIGNLLVPNDNYCKFED 179 | |||||||||||:|||||||||||:||||||||| |||||||||||||||||||||||| Db 121 HHMVDVIVTTTGGVEEDLIKCLAPTFKGDFSLPGAYLRSKGLNRIGNLLVPNDNYCKFED 180 Qy 180 WIIPILDQMLKEQLEENVLWTPSKVIQRLGREINDKNSYLYWAYKNNIPVFCPGLTDGSL 239 ||||| ||||||| ||||||||||:: |||:|||:::|||||||| |||||||||||||| Db 181 WIIPIFDQMLKEQKEENVLWTPSKLLARLGKEINNESSYLYWAYKMNIPVFCPGLTDGSL 240 Qy 240 GDMLYFHSFRNPGLIIDVIQDIRSMNGEAVHASPRKTGMIILGGGLPKHHICNANMMRNG 299 |||||||||| ||:|||:||||:||||||||:||||||||||||||||||||||||||| Db 241 GDMLYFHSFRTSGLVIDVVQDIRAMNGEAVHATPRKTGMIILGGGLPKHHICNANMMRNG 300 Qy 300 ADYAVFINTAQEFDGSDSGARPDEAVSWGKIRDSAKTVKVHCDATIAFPLLVAETFASRR 359 ||||||||| |||||||||||||||||||||| |||||||:|||||||||||||||||:| Db 301 ADYAVFINTGQEFDGSDSGARPDEAVSWGKIRGSAKTVKVYCDATIAFPLLVAETFASKR 360 Qy 360 EK 361 |: Db 361 EQ 362 The instant specification teaches that this 6-amino acid hypervariable region of the DHS protein is involved in the formation of subcellular protein complexes including substrate eIF-5A; small deletions of residues could result in less effective protein complex formations with reductions in senescence (page 8, lines 15-22). Plants with inhibited or reduced activity of DHS will have reduced active eIF-5A because there will be substantially less DHS protein to activate eIF-5A (page 15, lines 10-17). The specification teaches that these 6 amino acids have little conserved identity between plant species (page 8, lines 1-5). The specification teaches that in S. lycopersicum, the activity of the enzyme can be reduced by deleting (which has no corresponding residue in human DHS), E77, L78, P79, T80, or E81 (page 17, lines 11-15). The instant specification teaches multiple examples of gene editing technology to accomplish the deletion or insertion, such as an ARCUS engineered homing endonuclease to create a small in-frame deletion (page 17, lines 4-19) or an engineered transcription activator-like effector nuclease to create a double-stranded cleavage including the nucleotides surrounding the hypervariable region (page 17, line 21-page 18, line 8). The specification all teaches biallelic deletion mutations recovered in the hypervariable region of DHS1 in two tomato germplasms as a result of CRISPR methods (table 5, page 21, lines 5-9; page 19, line 16-page 20, line 7). The other examples provided by the specification, in soybeans (example 8), common bean (example 9), strawberry (example 10), pepper (example 11), zucchini (example 12), potato (example 13), rice (example 14), sorghum (example 15), and rose (example 16) are prophetic. These examples teach that the nucleic acid sequence for the hypervariable region will be used for targeting, and that the relevant region of the gene in the cultivar to be used must be sequenced first (for example, page 27, lines 7-19). A delayed senescence phenotype is not taught for the species other than tomato in the provided examples. The art teaches structural features and functions of DHS, although the 6 amino-acid hypervariable region and its function is not taught in the art. Wator-Wilk et al (2025) Structure. 33(2) (published 2/6/2025, hereafter Wator-Wilk) teaches that DHS is a tetramer with four identical active sites and is formed through the interaction of two monomers; the protein can form a dimer, which leads to a complete loss of activity (page 223, left column, paragraph 2). When eIF5A binds to the DHS tetramer, the remaining three entrances to active sites are sealed by a ball-and-chain regulatory motif which needs to be displaced upon binding to the protein substrate (Wator-Wilk page 223, right column, paragraph 1-2). The ball-and-chain motif has low sequence similarity between human and archaeal DHS (Wator-Wilk page 224, left column, paragraph 2). DHS remains catalytically active even when three active sites are inaccessible, suggesting that only one active site is sufficient to facilitate the hypusination of eIF5A (Wator-Wilk page 226, left column, paragraph 1). In Trypanosoma brucei, DHS is composed of two pairs of heterodimers (Wator-Wilk page 225, left column, paragraph 2). There was active-site complementation between two different paralogs of DHS in this species (Wator-Wilk page 222, right column, paragraph 1). DHS in plants is also known in the art. Feitelson et al US 2019/0203220 A1 (published 7/4/2019, hereafter Feitelson) teaches that down-regulation of DHS in plants is expected to generate plants with advantageous agronomic properties, such as resistance to stress, delayed senescence, and increased yield (paragraph [0011]). Feitelson teaches critical residues in the amino acid sequence of DHS include the residues from Glu323-Lys329, which are absolutely conserved in plant DHS sequences and is involved in the ability to catalyze the transfer of the butylamine group of spermidine to the Lys residue of eIF51 in humans (paragraph [0018]). Feitelson also teaches residues at 106, 238, 288, and 313 are conserved in plant DHS sequences and are involved in NAD+ binding (paragraph [0019]); tables 1 & 2). Feitelson describes mutations, substitutions, and deletions in human DHS protein (table 3); N-terminal deletions up to residue 48 or 97 lead to a complete loss of activity, while some residues can be mutated and rtain a measure of DHS activity (paragraph [0022]). Feitelson suggests that it may be possible to introduce a point mutation in an important residue that would reduce the activity of the modified gene but not eliminate it (paragraph [0022]), although Feitelson is silent as to the important of the six amino acid hypervariable region residues. Feitelson teaches a working example of gene editing DHS1 in rice plants using CRISPR-Cms1 with a guide RNA targeting either the intron or the active site (paragraphs [0070-0079]). Insertions in the intron-targeted plants were observed, and these plants had later flowering and increased chlorophyll (paragraph [0074-0075]), which might be considered delayed senescence if the plants are “staying green longer” (paragraph [0069]). A heterozygous rice plant for the active site target was recovered carrying a 10bp deletion with a frameshift leading to a stop codon and 41 fewer amino acids compared to the wild-type protein; the phenotype of these plants were shorter, had a higher tiller number, and had larger seed and darker green appearance (paragraph [0078]). Feitelson does teach a heterozygous Brassica napus plant comprising a 1 bp deletion adjacent to a targeted sequence resulting in a truncated DHS protein and delayed leaf senescence and dwarf plants (paragraphs [0062-0069]), so a senescence phenotype has been observed in plants comprising mutated DHS. In Arabidopsis, a DHS mutant possess a lethal female gametophyte and cannot be a homozygous diploid (Palfi et al (2021) Plants. 10(7) published 6/22/2021, hereafter Palfi. page 4, paragraph 2). However, there may be a functional difference in DHS between different plant species; leaf-specific suppression of DHS in Arabidopsis resulted in growth enhancement without any negative pleiotropic effects in one study, although pleiotropic effects were taught in the same plants in an earlier study, while suppression of DHS in petunia led to abnormal chloroplast development and delated leaf senescence (Palfi page 4, paragraph 2). In tomato plants, there are four SleIF5As, and only one SlDHS gene can take part in the posttranslational activation of all SleIF5A isoforms; however, there is no evidence regarding the level of functional redundancy in senescence processes in plants (Palfi page 6, paragraph 5). One of ordinary skill in the art would not know, based on the prior art and the instant specification, whether plants that have multiple DHS genes may exhibit senescence as a result of any deletion or insertion in the hypervariable region of the instant invention in any copy of DHS. This is further reinforced by the teachings of the art of uncertainty regarding the function of DHS paralogs. While the main activity of DHS is the activation of eIF5A, plant DHS are known to be promiscuous with regard to the utilization of aminobutyl donors and acceptors, such as transferring the aminobutyl moiety from spermidine to putrescine to yield homospermidine, an important precursor of pyrrolizidine alkaloids (Kaltenegger et al (2021) FEBS Open Bio. 11: 10-25 (Published 11/28/2020, hereafter Kaltenegger) page 11, left column, paragraph 3-right column, paragraph 1). DHS is highly promiscuous and can catalyze various reactions, including some of unknown biological relevance (abstract). Duplicates of genes that encode homomeric proteins like DHS are predicted to interfere at the protein level by forming paralogous heteromers (page 12, left column, paragraph 1). Repeated independent duplications of DHS in plant lineages followed by functional divergence gave rise to HSS, and the plant Lolium perenne comprises two DHS paralogs (page 21, left column, paragraph 1). However, the biochemical characteristics of newly identified DHS paralogs are unknown (page 21, left column, paragraph 1). Although mutations in DHS in plants are known, insertion or deletion mutations specifically in the recited hypervariable region of the instant specification are not taught in the art. Adams et al US 2009/0100536 A1 (published 4/16/2009, hereafter Adams) teaches a polypeptide sequence (Adams SEQ ID NO: 489) which is a Glycine max deoxyhypusine synthase (table 2) and has 99% sequence similarity to 94% of instant SEQ ID NO: 45. See alignment below. Score Expect Method Identities Positives Gaps 765 bits(1976) 0.0 Compositional matrix adjust. 366/368(99%) 367/368(99%) 0/368(0%) Query 26 MGEAKGKEEEVLASVHSTVFKESESLDGKCVKIEGYDFNRGVNYPLLLSSMVTTGFQASN 85 MGEAKGKEEEVLASVHSTVFKESESLDGKCVKIEGYDFNRGVNYPLLLSSMVTTGFQASN Sbjct 1 MGEAKGKEEEVLASVHSTVFKESESLDGKCVKIEGYDFNRGVNYPLLLSSMVTTGFQASN 60 Query 86 LGDAIQVVNQMLDWRLVDEPVAEDCSDQERDLEYRKSVTCKVFLGFTSNLISSGVRDTVR 145 LGDAIQVVNQMLDWRLVDEPVAEDCSDQERDLEYRKSVTCKVFLGFTSNLISSGVRDTVR Sbjct 61 LGDAIQVVNQMLDWRLVDEPVAEDCSDQERDLEYRKSVTCKVFLGFTSNLISSGVRDTVR 120 Query 146 FLLQHRMVDVVVTTTGGIEEDLIKCLAPTYKGDFSLPGAYLRSKGLNRIGNLLVPNDNYC 205 FLLQHRMVDVVVTTTGGIEEDLIKCLAPTYKGDFSLPGAYLRSKGLNRIGNLLVPNDNYC Sbjct 121 FLLQHRMVDVVVTTTGGIEEDLIKCLAPTYKGDFSLPGAYLRSKGLNRIGNLLVPNDNYC 180 Query 206 KFEDWIIPIFDQMLGEQNNENVLWTPSKLIARLGKEIKNESSYLYWAYKNNIPVFCPGLT 265 KFEDWIIPIFDQMLGEQNNENVLWTPSKLIARLGKEIKNESSYLYWAYKNNIPVFCPGLT Sbjct 181 KFEDWIIPIFDQMLGEQNNENVLWTPSKLIARLGKEIKNESSYLYWAYKNNIPVFCPGLT 240 Query 266 DGSLGDMLYFHSFRNPGLIVDIVQDIRAMNGEAVHASPRKTGMIILGGGLPKHHICNANM 325 DGSLGDMLYFHSFRNPGLIVDIVQDIRAMNGEAVHASPRKTGMIILGGGLPKHHICNANM Sbjct 241 DGSLGDMLYFHSFRNPGLIVDIVQDIRAMNGEAVHASPRKTGMIILGGGLPKHHICNANM 300 Query 326 MRNGADYAVFINTAQEYDGSDSGARPDEAVSWGKIRDSAQTVKVHCDATIAFPLLVAETF 385 MRNGADYAVFINTAQEYDGSDSGARPDEAVSWGKIRDSAQTVKVHCDATIAFPLLVAETF Sbjct 301 MRNGADYAVFINTAQEYDGSDSGARPDEAVSWGKIRDSAQTVKVHCDATIAFPLLVAETF 360 Query 386 ASRVKPHH 393 AS+VKP H Sbjct 361 ASKVKPPH 368 Adams teaches soybean transformed with each gene in table 2, and that progeny transgenic plants and seed are screened for enhanced water use efficiency, cold tolerance, nitrogen use efficiency, seed protein, and seed oil and for increased yield (paragraphs [0082-0084]). Adams teaches that a nucleotide sequence encoding a maize DHS polypeptide sequence (Adams SEQ ID NO: 214), when transformed into maize plants, improved growth in high nitrogen (table 9) in limited nitrogen (table 10), and under cold stress (table 16), improved water use efficiency (table 15), improved yield (table 14), improved protein content in seeds (table 19) and improved oil content in seeds (table 20). Adams teaches a Glycine max amino acid sequence, Adams SEQ ID NO: 3827, with an insertion relative to instant SEQ ID NO: 45 in the hypervariable region of the instant application and 90% sequence similarity. See alignment below; hypervariable region is in bold. Adams teaches this is a homologous protein to the proteins used in the transgenic assays (paragraph [0018]). Adams suggests plant cell nuclei with stably integrated, recombinant DNA comprising a promoter operably linked to a homolog such as Adams SEQ ID NO: 3827 (claim 1(c)). Although Adams suggests that a homolog of a Glycine max DHS gene comprising an insertion in the hypervariable region might be able to provide the same benefits as a DHS gene expressed as a transgene, this is not demonstrated in a working example, and the actual effect on senescence of such a coding sequence is not taught. AWW69160 ID AWW69160 standard; protein; 387 AA. XX AC AWW69160; XX DT 12-NOV-2009 (first entry) XX DE Glycine max protein, SEQ ID 3827. XX KW Plant breeding; crop improvement; transgenic plant; plant. XX OS Glycine max. XX CC PN US2009100536-A1. XX CC PD 16-APR-2009. XX CC PF 18-JUL-2007; 2007US-00879785. XX PR 04-DEC-2001; 2001US-0337358P. PR 04-DEC-2002; 2002US-00310154. PR 13-DEC-2002; 2002US-00319154. XX CC PA (MONS ) MONSANTO CO. XX CC PI Adams T, Agarwa AK, Ahrens J, Ball JA, Basra A, Bell E; CC PI Bradshaw TL, Chomet PS, Crowley JH, Deikman J, Deng M, Donnarummo M; CC PI Duff KFZ, Duff S, Edgerton MD, Gopalan B, Heal M, He X, Hinchey BS; CC PI Huang S, Johnson RG, Jung V, Kretzmer K, Laccetti LB, Lai CQ; CC PI Lee GJ, Leland TJ, Liu J, Lu B, Luethy MH, Lund AA, Madson LL; CC PI Malloy KP, Mcnabnay MP, Padmavati M, Ruff T, Start W, Tennessen D; CC PI Urwin CP, Vaduva G, Vidya KR, Voss ST, Voyles C, Xin Z, Xu N; CC PI Xu Q, Zhang Q, Zhao Y, Zhou L; XX DR WPI; 2009-H07992/29. XX CC PT Novel plant cell nucleus stably integrated with recombinant DNA having CC PT promoter operably linked to DNA encoding amino acid sequence having Pfam CC PT domain module, useful for generating transgenic plant cell, seed or plant CC PT e.g. corn. XX CC PS Claim 2; SEQ ID NO 3827; 0pp; English. XX CC The present invention relates to a novel plant cell nucleus which is CC stably integrated with recombinant DNA having a functional promoter CC operably linked to a DNA encoding a protein which comprises a Pfam domain CC module. The plant cell nucleus of the invention can be used to generate CC transgenic plants with an enhanced trait including enhanced water use CC efficiency, enhanced cold tolerance, increased yield, enhanced nitrogen CC use efficiency, enhanced seed protein and/or enhanced seed oil. The CC transgenic plant cell can be a crop plant, transgenic pollen grain or CC transgenic seed of corn, soybean, cotton, canola, alfalfa, wheat or rice CC plant. The present sequence is a protein which imparts an enhanced trait CC in plant cells. XX SQ Sequence 387 AA; Query Match 90.8%; Score 1878.5; Length 387; Best Local Similarity 98.6%; Matches 359; Conservative 1; Mismatches 3; Indels 1; Gaps 1; Qy 26 MGEAKGKEEEVLASVHSTVFKESESLDGKCVKIEGYDFNRGVNYPLLLSSMVTTGFQASN 85 ||||||||||||| |||||||||||||||||||||||||||||||||||||||||||||| Db 1 MGEAKGKEEEVLAPVHSTVFKESESLDGKCVKIEGYDFNRGVNYPLLLSSMVTTGFQASN 60 Qy 86 LGDAIQVVNQMLDWRLVDE-PVAEDCSDQERDLEYRKSVTCKVFLGFTSNLISSGVRDTV 144 ||||||||||||||||||| ||||||||||||||||||||||||||||||||||||||| Db 61 LGDAIQVVNQMLDWRLVDEXXVAEDCSDQERDLEYRKSVTCKVFLGFTSNLISSGVRDTV 120 Qy 145 RFLLQHRMVDVVVTTTGGIEEDLIKCLAPTYKGDFSLPGAYLRSKGLNRIGNLLVPNDNY 204 |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||| Db 121 RFLLQHRMVDVVVTTTGGIEEDLIKCLAPTYKGDFSLPGAYLRSKGLNRIGNLLVPNDNY 180 Qy 205 CKFEDWIIPIFDQMLGEQNNENVLWTPSKLIARLGKEIKNESSYLYWAYKNNIPVFCPGL 264 |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||| Db 181 CKFEDWIIPIFDQMLGEQNNENVLWTPSKLIARLGKEIKNESSYLYWAYKNNIPVFCPGL 240 Qy 265 TDGSLGDMLYFHSFRNPGLIVDIVQDIRAMNGEAVHASPRKTGMIILGGGLPKHHICNAN 324 |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||| Db 241 TDGSLGDMLYFHSFRNPGLIVDIVQDIRAMNGEAVHASPRKTGMIILGGGLPKHHICNAN 300 Qy 325 MMRNGADYAVFINTAQEYDGSDSGARPDEAVSWGKIRDSAQTVKVHCDATIAFPLLVAET 384 |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||| Db 301 MMRNGADYAVFINTAQEYDGSDSGARPDEAVSWGKIRDSAQTVKVHCDATIAFPLLVAET 360 Qy 385 FASR 388 || : Db 361 FAXK 364 Thus, although one of skill in the art would be able to sequence a DHS gene, design a gRNA to target the recited region, and recover mutations, some of which could be expected to comprise insertions and/or deletions, one of skill in the art would not necessarily know that any such insertion or mutation in any DHS paralog has a reduced DHS activity and delayed senescence in any species of plant. Rather, one of ordinary skill in the art would be required to screen plants, including those with multiple DHS genes, to determine which mutations, in which DHS genes, reduce a DHS activity, and of those with a reduced activity, which ones provide delayed senescence. The uncertainty of DHS redundancy, the multiple DHS activities known in plants, and the lack of teachings in the art regarding this hypervariable region would require an undue burden of experimentation for an ordinary practitioner in the art to practice the method and generate the plants over the full scope of the claimed invention. Claim 25, which requires that the plant is Solanum lycopersicum and that the DHS comprises the amino acid sequence of SEQ ID NO: 87, is not included in the rejection because one of skill in the art would be enabled to make and test insertions and deletions in this species, as taught in the specification. Conclusion No claims are allowed. Any inquiry concerning this communication or earlier communications from the examiner should be directed to Victoria L DeLeo whose telephone number is (703)756-5998. The examiner can normally be reached M-F 8:00am-4pm EDT. Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice. If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Bratislav Stankovic can be reached at (571) 270-0305. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300. Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000. /VICTORIA L DELEO/Examiner, Art Unit 1662 /Anne Kubelik/Primary Examiner, Art Unit 1663