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 .
Claim Status
Claims 1-17 are pending.
Claims 1-17 are examined on the merits.
Claim Objections
Claim 1 is objected to because of the following informality: the duplicate Markush list includes "a 7 base pair deletion from nucleotide 1370 to nucleotide 1376, as compared to reference sequence SEQ ID NO:4" (p4 and p5). Appropriate correction is required.
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.
Claims 1-6, and 8-15 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 is rejected as being indefinite because the metes and bounds of the term “an endogenous TFL1b gene” and thus “a promoter of an endogenous TFL1b gene” are unclear. The claim does not provide objective boundaries for what sequences constitute the claimed TFL1b gene, it is unclear whether “TFL1b” is being defied by (1) a particular gene/locus in soybean, (2) a nucleotide sequence (coding region and/or genomic sequence), (3) a protein identity or sequence, and/or (4) a function (e.g., a gene encoding a TFL1-like protein involved in flowering). The designation “TFL1b” is a name-based label that, without an express sequence /locus definition in the claim, can reasonably refer to different genes/alleles/paralogs/orthologs within soybean germplasm and across references, such that a person of ordinary skill in the art cannot determine with reasonable certainty which endogenous gene is required by the claim.
Further, the claim defines the promoter modification using nucleotide positions “as compared to reference sequence SEQ ID NO:4” (e. g. deletions from nucleotide 1128-1131. 1367-1403, etc.), but the claim does not clearly state whether SEQ ID NO: 4 is the promoter sequence of the claimed endogenous TFL1b gene, nor does it recite any sequence identity/structural requirement tying the “promoter of an endogenous TFL1b gene” to SEQID NO:4 (or explaining how to determine corresponding nucleotide positions in different allelic promoter sequences). Because endogenous promoter sequences commonly vary among soybean lines, a person of ordinary skill in the art would not be able to determine, with reasonable certainty, whether a given plant’s endogenous promoter sequence corresponds to SEQ ID NO:4 such that the recited “nucleotide 1128-1131”(etc.) are meaningfully identified and whether the plant falls within the scope of the claim. Accordingly, the scope of the claimed subject matter is indefinite.
Claim 2 does not resolve the ambiguity of claim 1, because it characterizes the “TFL1b gene” by a broad protein identity range to SEQ ID NO:2 as low as 70%, which encompasses a broad range of related sequences that could corresponds to different genes within the TFL1b/Dt1 family, including genes that are highly homologous yet functionally distinct (e. g., TFL1a versus TFL1b). Furthermore, soybean contains multiple TFL1-like genes, TFL1a and TFL1b are highly homologous (Baohui Liu et. al., The soybean stem growth habit gene Dt1 is an ortholog of Arabidopsis TERMINAL FLOWER1, Plant Physiol. 2010 May;153(1):198-210, 2010 Mar 10), but differ in function (Liu 2010, p205, discussion), and promoter region (see alignment below). The “TFL1b gene” by a broad protein identity range to SEQ ID NO:2 as low as 70% may correspond to different TFL1 family members having different promoters. Because different loci/genes would have different promoter sequences, a person of ordinary skill in the art cannot determine with reasonable certainty which endogenous gene and which promoter are required by the claims, nor how the promoter modifications defined by coordinate relative to SEQ ID NO:4 correspond to the promoter of the claimed “endogenous TFL1b gene” across the full scope. thus, the relationship between the promoter sequence of SEQ ID NO:4 and the “endogenous TFL1b gene” recited in claim 1 still remains unclear.
Dependent claims 3-6 and 8-15 are included in this rejection because they do not include additional limitations to resolve the ambiguity.
CLUSTAL O(1.2.4) multiple sequence alignment
TFL1b MAKMPLEPLIVGRVIGEVLDSFTTSTKMTVSYNKKQVYNGHELFPSTVNTKPKVEIEGGD 60
TFL1a MARMPLEPLIVGRVIGEVLDSFTTSTKMIVSYNKNQVYNGHELFPSTVNTKPKVEIEGGD 60
**:************************* *****:*************************
TFL1b MRSFFTLIMTDPDVPGPSDPYLREHLHWIVTDIPGTTDATFGKELVSYEVPKPNIGIHRF 120
TFL1a MRSFFTLIMTDPDVPGPSDPYLREHLHWIVTDIPGTTDATFGKELVSYEIPKPNIGIHRF 120
*************************************************:**********
TFL1b VFVLFKQKRRQCVTPPTSRDHFNTRKFAAENDLGLPVAAVYFNAQWETAARRR 173
TFL1a VFVLFKQKRRQCVTPPTSRDHFNTRKFAAENDLALPVAAVYFNAQRETAARRR 173
*********************************.*********** *******
CLUSTAL O(1.2.4) multiple sequence alignment for promoter region of TFL1a/TFL1a (AB511820 and AB511821)
TFL1a ------------------------------------------------------------ 0
TFL1b caattttaaattatatagttatacacaaataaacaaaatataaaaaaaacaaaaatcaaa 60
TFL1a ------------------------------------------------------------ 0
TFL1b tcatgtgaagattttcaaaaaaatacatgacttttagaaaaaatcacaaaacttgcatgt 120
TFL1a ------------------------------------------------------------ 0
TFL1b atgagagaaagagacgagggttaattaatttaacttcttcaaaattttctttttattttt 180
TFL1a ------------------------------------------------------------ 0
TFL1b tctgtatgtatgcatatattatttgaaaatttaattgaaatacaatgataatataaggat 240
TFL1a ------------------------------------------------------------ 0
TFL1b tttttatactgttgtttaatcataaaacatcatatagtgaaaatttgttaaattttatac 300
TFL1a ------------------------------------------------------------ 0
TFL1b aataattattttaaaaaattatcttaaaaaattactttaaaaaaatacatttttggtaca 360
TFL1a ------------------------------------------------------------ 0
TFL1b tataaaatttatactaacagtatataaaaattaagctaaatataaaaagtatatttaaaa 420
TFL1a ------------------------------------------------------------ 0
TFL1b aaattatgttgtatataataccaatatgaccaacgaatattccatgcttaatcggcatca 480
TFL1a ------------------------------------------------------------ 0
TFL1b ctaacttttaatcaagttaaacgaagaaagtaggaactaaaagataaacatgagcaactt 540
TFL1a ------------------------------------------------------------ 0
TFL1b agcattttaattataatgatattggaagtttaggaaagaaaacctaaaatatccaaattt 600
TFL1a ------------------------------------------------------------ 0
TFL1b agaaatatgaatatcaacttataccaaaataagcaactaacagtttcagaccaacaaatt 660
TFL1a ------------------------------------------------------------ 0
TFL1b ttagaatcatcatctccatataccacataattaataattaagtctcaaattgaaataata 720
TFL1a -----------------------------ctgcaatttatttatgcaataatttgtctat 31
TFL1b ttatattttcaaagtgaaacgataccaacctgcaatttatttatgcaataaacggtctat 780
********************** ******
TFL1a atattccactttcaaatcataatataaaaaaatcaatgtaacgaatgtctgcttaattaa 91
TFL1b atattccactttcaaatcataatataaaaaaatcaatataacgaatgtctgcttaattaa 840
************************************* **********************
TFL1a ttatacgtacgaaacta----tgccaccgccaattagggtttttttattttattttctgg 147
TFL1b ttatacgtactaattaaactatgccaccaccaattaggg-----cttttttattttctgg 895
********** ** * ******* ********** * *************
TFL1a aactagaatttgtgatctttaaagcaagggatgcgcatggatgcacacacgtcaacccct 207
TFL1b aactagaatttgtgatcttttaagcacgggatgtgcatgtacac----acacctcaaccc 951
******************** ***** ****** ***** * * ** * **
TFL1a ttttttctcctacaaaagaatgcaaacagaacttgaataacataaaatttcacatggaga 267
TFL1b ctttttctccaacaaaagaatgcaaacagaacttgaataacataaaatttcacatgcaga 1011
********* ********************************************* ***
TFL1a gatgaggagggcaaaatggtttcatgataaagg---tccagcttcctcttttaccaatct 324
TFL1b gatgaagagggcaaaatggtttcttgataaagaaggtccaccttcctcttttaccaatat 1071
***** ***************** ******** **** ***************** *
TFL1a tggaactttggtctgaatttaagagttaagtttttattctgaaatgacctcttaggcact 384
TFL1b -tggaacttggtctgcatctaagagttaagtttttattctgaaatgacctcttaggcact 1130
* * ******** ** *****************************************
TFL1a cacacacttgtactctataggcgtgtctagccttgagcatttgtggtggagttcctttaa 444
TFL1b caaacgcttatactccataggcgtgtctagccttgagaatttgtggtggagttcctttaa 1190
** ** *** ***** ********************* **********************
TFL1a ttatgccatctttatcacgaatcttccaacacaattaaaagaaagactatggctctgagg 504
TFL1b ttatgccctcttcatcaggaatcttccaacaaaattaaaagaaagactatggctaaggga 1250
******* **** **** ************* ********************** * *
TFL1a gaaaatggagaatcacgcacccacaaacagaaatgttgcacattgtgtgcatgcaattaa 564
TFL1b --aaatggagaatcacgcacgcacaaacag-aaatttgcatagtgtgtgcaaacaattaa 1307
****************** ********* ** ***** * ******** *******
TFL1a ggaaggttctctacgcaaggcatacaacaaaaatttctgcacgtccatggtactaaga-- 622
TFL1b ggaaggttctctacgcaaggcatacaacagaaatttctgcacgtccatggtactagtagt 1367
***************************** ************************* *
TFL1a ----------gaatgctaaaatgtgggtgttcaagaaacgaagcacttcctagagttgtt 672
TFL1b atactaaagagaatgctaaaatgtgggtgttcaagaaacgaagcatttcctagaattttt 1427
*********************************** ******** ** **
TFL1a cccttttatgcttttagtttaataatgaaacggcagaagcttgacaacttgcaaagggag 732
TFL1b gccttttatgcttttagtttaataataaaacggcagaagcttgacaacttgcaaagggga 1487
************************* *******************************
TFL1a aggaccca-------------catacacatgatgcttttttttacctgatcgagctgatg 779
TFL1b gggaggacccacacacacaggattacacatgatgctttttttgacctgatcgagctgatg 1547
*** ******************* *****************
TFL1a ggaagaaaaaaatgggaagtttgtattttttaatattttta----caatttttttagaac 835
TFL1b ggagagaaaaaactgggaaattttatatttttaaatattttttacaaattttttcagaac 1607
*** ****** ** * ** *** **** * ** *** ******** *****
TFL1a tgggatggatgggatggaattaaaa-agaagtacactgtaaaagggttgttattagaata 894
TFL1b taggatggg----atagaaataaataaaaaaagtacacaataagagttgctaatagaata 1663
* ****** ** *** **** * ** ** * *** **** ** *******
TFL1a aaaaaaaattgctttattggatggaaatatccatgcatccattcctgtgtacagccaatc 954
TFL1b a--aaaaatttctttatt----ggatgagaaatatccatgcatccggtgtacagccaatc 1717
* ******* ******* *** * *** **************
TFL1a aattaagaaccgattctaatggccccattctgtgtttatgaccaggggagaga------- 1007
TFL1b aattaagaaccgattccaagggccccattctgtgtttatgagcagcggagagagaaagag 1777
**************** ** ********************* *** *******
TFL1a ---ggaaaaggggaaaggataatttgtcacatatcatgactaatgtgatttcagcacaaa 1064
TFL1b agtggaaaagtggaaaggataactcgtcacatatcatgactaatgtgatttcagcacaaa 1837
******* *********** * ***********************************
TFL1a catcccagacccaaaatcaattgttgagatgagccaaattaagagataatatattagggg 1124
TFL1b catcccagacccaaaggcaattgttgagatgagccgaattaagagacaatatattagtag 1897
*************** ****************** ********** ********** *
TFL1a tagtgggaggaccttcacggatgcacttgatttatttgtaggcttttttcattttttgtt 1184
TFL1b tagtgggaggacctgcacggatgcacttgatttatttctagccttttttcattttttgtt 1957
************** ********************** *** ******************
TFL1a tttcctgtagaacatgtttttt---tctctaaatctaaagaatgtgttgcaacatctttt 1241
TFL1b tttcctatagaacttgtattttctctagctaaatctaaattatgtattgcaacatatttt 2017
****** ****** *** **** * *********** **** ********* ****
TFL1a aaaggaggggggaaaaacactctaccttgaa---agacactttttaagacattataaata 1298
TFL1b aaaggaggggcaaaacacactcgatctaccttcaaagaaacttttaagacattataaata 2077
********** *** ****** * ** * * *******************
TFL1a tagagctcacactccctttcctcagcaggattcatcataacaattgtattcctcccttcc 1358
TFL1b tggagcccacactcccttcttcctcagtaag---atcatcaacttgtattcctcacttcc 2134
* **** *********** * * * *********** *****
TFL1a ttagctccactacctcttctctcttcctccttgttccttcctcttaca 1406
TFL1b ttagctcctcctcttccttgttcctcctcttaca-------------- 2168
******** * * ** * ** ***** *
Claim 1 is rejected as indefinite for the recitation “the modification is selected from the group consisting of………and combinations of any thereof”. Many listed deletions overlap with each other. If “combine “two overlapping deletions, the real allele may become one merged deletion, it’s unclear whether that merged deletion is still a “combination”, or must match two discrete listed members.
Dependent claims 2-6, and 8-15 are included in this rejection because they do not include additional limitations to resolve the ambiguity.
Claim 14 is rejected as being indefinite for the recitation “plant exhibits substantially the same..”. “substantially” is a term of degree that lacks objective boundaries in the claim; the claim does not specify what range or tolerance qualifies as “substantially the same”, nor the conditions/measurement used to determine, such that one of ordinary skill in the art cannot determine the metes and bounds with reasonable certainty.
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.
Written Descriptions
Claims 1-6, 8-11 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention.
The Federal Circuit has clarified the application of the written description requirement. The court stated that a written description of an invention "requires a precise definition, such as by structure, formula, [or] chemical name, of the claimed subject matter sufficient to distinguish it from other materials". University of California v. Eli Lilly and Co., 119 F.3d 1559, 1568; 43 USPQ2d 1398, 1406 (Fed. Cir. 1997). The court also concluded that "naming a type of material generally known to exist, in the absence of knowledge as to what that material consists of, is not description of that material". Id. Further, the court held that to adequately describe a claimed genus, Patent Owner must describe a representative number of the species of the claimed genus, and that one of skill in the art should be able to "visualize or recognize the identity of the members of the genus". Id.
The claims are rejected for lacking adequate written description support regarding the broad scope of the following.
Claim 1 regards the “a promoter of an endogenous TFL1b gene” and “endogenous TFL1b gene”.
Claim 2 regards “at least 70% … sequence identity to SEQ ID NO:2”.
“TFL1b gene” refers to a gene in the FT/TFL1 (PEBP) family that encodes a TFL1-like (floral repression/indeterminacy-associated) protein; the “b” designation indicates a particular paralog in the species and does not, by itself, uniquely identify a single genomic locus absent a sequence/locus definition. (Lan Wu et. al., Identification and Characterization of the FLOWERING LOCUS T/TERMINAL FLOWER 1 Gene Family in Petunia, DNA and Cell Biology Vol. 38, No. 9, 2019) and (Lillian Magalhães Azevedo et. al., The Role of FT/TFL1 Clades and Their Hormonal Interactions to Modulate Plant Architecture and Flowering Time in Perennial Crops, Plants, 2025, 14(6), 923).
Claim 1 covers any “endogenous TFL1b gene” in soybean and its promoter, where the promoter has one of the listed deletions (defined by coordinates relative to SEQ ID:4) and the modification reduces TFL1b expression/activity, and claim 2 expends “ TFL1b” gene to include genes encoding proteins having as low as low as 70% identity to SEQ ID NO:2. Soybean has multiple TFL1 homologs, they are highly conservative (Liu(2010), example from above alignment), so the term “TFL1b gene” is not inherently single-locus.
The specification describes a particular soybean determinacy locus identified as GmTFL1b/Dt1, provides a specific coding sequence (SEQ ID NO:1), protein sequence (SEQ ID NO:2), cDNA (SEQ ID NO:3), and a specific upstream promoter region (SEQ ID NO:4) for targeted editing, and describes example guide RNAs and example promoter edits within that referenced promoter sequence, with resulting expression measurements in selected edited lines.
The specification does not reasonably convey possession of the full scope of the claimed “endogenous TFL1b gene” and “promoter of an endogenous TFL1b” as claimed.
The claims cover a broad “TFL1b”gene genus that is not adequately described. Claim 2 extends the “TFL1b gene” to genes encoding proteins having as low as 70% identity to SEQ ID NO:2. A protein identity threshold of 70% is broad enough to encompass a wide range of relate sequences (one example of blast SEQ ID NO:2 shows below), such as GlymaFiskIII.16G173500.1 (the Fiskeby III gene model corresponding to the Glyma.16G173500 locus name used in Wm82-based papers) is annotated as a SLAC/SLAH-family S-type anion channel, which share 74% identity of SEQ ID NO:2. The example also includes other members of the soybean PEBP/TFL1-like gene family (such as TFL1a, c, and d), which are known to include multiple homologs in soybean, Wang (Lingshuang Wang et. al., Two soybean homologues of TERMINAL FLOWER 1 control flowering time under long day conditions, The Crop Journal, Volume 11, Issue 3, June 2023, Pages 704-712). The alignment result of TFL1d with SEQ ID NO: 2 is also listed below. Moreover, as mentioned before, soybean contains closely related homologs such as GMTFL1a, which were reported to have high amino acid identity yet different transcriptional profiles Liu(2010).
Thus, sequence similarity alone, especially at the breadth recited, does not uniquely identify the claimed “TFL1b gene” as the specific Dt1/GmTFL1b locus.
The promoter scope is not commensurate with what is disclosed. Claim 1 requires a modification in the “promoter of an endogenous TRL1b gene”, yet the recited modifications are defined by coordinates relative to SEQ ID NO:4. If claim 2 encompasses additional “TFL1b genes” meeting the broad identity, threshold (including other homologs or alleles), those different lock would be expected to have different upstream regulatory sequences (i. e., different promoters), and the coordinate-based deletions defined relative to SEQ ID NO:4 would not necessarily correspond to the promoter of those other encompassed genes. The specification provides disclosure directed to editing one particular promoter sequence (SE ID NO:4) but does not describe representative promoter sequences, alignment/mapping rules, or distinguishing structural features sufficient to identify and delimit “the promoter of an endogenous TFL1b gene” across the breadth of genes encompassed by claim 2.
Liu(2010) (p198) point out that GmTFL1a vs GmTFL1b differ in expression profiles despite high similarity, these proves the “similar gene” not same as regulation/promoter behavior. Even for the same gene (GmTFL1b/Dt1), the upstream promoter sequence can vary among references/alleles (e.g., the promoter region shown in Liu(2010) does not perfectly align with SEQ ID NO:4), demonstrating that “the TFL1b promoter” is not a single invariant sequence. (see promoter alignment below).
For a genus claim defined by sequence identity and functional outcome (reduced expression/activity), the specification must provide either a representative number of species across the genus, or common structural features correlating to the function. Here, the disclosure is directed to a specific soybean Dt1/GmTFL1b locus and a specific promoter region (/seq ID NO:4) with example edits,. The specification does not describe which sequence features distinguish “TFL1b” from other closely related PEBP family members, nor does it provide guidance establishing that proteins down to 70% identity to SEQ ID NO:2 necessarily correspond to the same endogenous gene (TFL1b) or will be regulated by eh promoter sequence of SEQ ID NO:4. In general, PEBP-family proteins can be homologous yet have distinct biological roles (e.g., FT versus TFL1), illustrating that sequence similarity alone may be insufficient to define the claimed gene/function boundary. Wickland (Daniel P. Wickland et. al., The FLOWERING LOCUS T/TERMINAL FLOWER 1 Gene Family: Functional Evolution and Molecular Mechanisms, Molecular Plant, Volume 8, Issue 7, 6 July 2015, Pages 983-997).
Furthermore, claim 1’s promoter modification are anchored to SEQ ID NO:4, implying a specific promoter reference, while claim 2 defines the “TFL1b gene” by a broad protein identity range that can encompass other homologs and variants. The specification does not describe promoter sequences and mapping rules sufficient to establish that SEQ ID NO:4 is the promoter for each encompassed “TFL1b gene” withing claim 2’s scope. Accordingly, the specification does not reasonably convey possession of the full claimed subject matter.
Claims 2-6, and 8-11 depend from claim 1, are included. The limitations do not remedy the lack of written description support for the full scope of the claimed “endogenous TFL1b gene” and its promoter.
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TFL1bpromoter ------------------caattttaaattatatagttatacacaaataaacaaaatata 42
SEQIDNO4 attgaattttttaagaaacaattttaaattatatagttatacataaataaacaaaataca 60
************************* ************** *
TFL1bpromoter aaaaaaacaaaaatcaaatcatgtgaagattttcaaaaaaatacatgacttttagaaaaa 102
SEQIDNO4 aaaaaaacaaaaatcaaatcatgtgaagattttcaaaaaaatacatgacttttagaaaaa 120
************************************************************
TFL1bpromoter atcacaaaacttgcatgtatgagagaaagagacgagggttaattaatttaacttcttcaa 162
SEQIDNO4 atcacaaaacttgcatgtatgagagaaagagacgagggttaattaatttaacttcttcaa 180
************************************************************
TFL1bpromoter aattttctttttattttttctgtatgtatgcatatattatttgaaaatttaattgaaata 222
SEQIDNO4 aattttctttttattttttctgtatgtatgcatatattatttgaaaatttaattgaaata 240
************************************************************
TFL1bpromoter caatgataatataaggattttttatactgttgtttaatcataaaacatcatatagtgaaa 282
SEQIDNO4 cactgataatataaggattttttatactgttgtttaatcataaaacatcatatagtgaaa 300
** *********************************************************
TFL1bpromoter atttgttaaattttatacaataattattttaaaaaattatcttaaaaaattactttaaaa 342
SEQIDNO4 atttgttaaattttatacaataattattttaaaaaattatcttaaaaaattactttaaaa 360
************************************************************
TFL1bpromoter aaatacatttttggtacatataaaatttatactaacagtatataaaaattaagctaaata 402
SEQIDNO4 aaatacatttttggtacatataaaatttatactaacagtatataaaaattaagctaaata 420
************************************************************
TFL1bpromoter taaaaagtatatttaaaaaaattatgttgtatataataccaatatgaccaacgaatattc 462
SEQIDNO4 taaaaagtatatttaaaaaaattatgttgtatataataccaatatgaccaacgaatattc 480
************************************************************
TFL1bpromoter catgcttaatcggcatcactaacttttaatcaagttaaacgaagaaagtaggaactaaaa 522
SEQIDNO4 catgcttaatcggcatcactaacttttaatcaagttaaacgaagaaagtaggaactaaaa 540
************************************************************
TFL1bpromoter gataaacatgagcaacttagcattttaattataatgatattggaagtttaggaaagaaaa 582
SEQIDNO4 gataaacatgagcaacttagcattttaattataatgatattggaagtttaggaaagaaaa 600
************************************************************
TFL1bpromoter cctaaaatatccaaatttagaaatatgaatatcaacttataccaaaataagcaactaaca 642
SEQIDNO4 cctaaaatatccaaatttagaaatatgaatatcaacttataccaaaataagcaactaaca 660
************************************************************
TFL1bpromoter gtttcagaccaacaaattttagaatcatcatctccatataccacataattaataattaag 702
SEQIDNO4 gtttcagaccaacaaattttaaaatcatcatctccatatgccacataattaataattaag 720
********************* ***************** ********************
TFL1bpromoter tctcaaattgaaataatattatattttcaaagtgaaacgataccaacctgcaatttattt 762
SEQIDNO4 tctcaaattgaaataatattatattttcacagtgaaacgataccaacctgcaatttattt 780
***************************** ******************************
TFL1bpromoter atgcaataaacggtctatatattccactttcaaatcataatataaaaaaatcaatataac 822
SEQIDNO4 atgcaataaacggtctatatattccactttcaaatcataatataaaaaaatcaatataac 840
************************************************************
TFL1bpromoter gaatgtctgcttaattaattatacgtactaattaaactatgccaccaccaattagggctt 882
SEQIDNO4 gaatgtctgcttaattaattatacgtactaattaaactatgccaccaccaattagggctt 900
************************************************************
TFL1bpromoter ttttattttctggaactagaatttgtgatcttttaagcacgggatgtgcatgtacacaca 942
SEQIDNO4 ttttattttctggaactagaatttgtgatcttttaagcacgggatgtgcatgtacacaca 960
************************************************************
TFL1bpromoter cctcaacccctttttctccaacaaaagaatgcaaacagaacttgaataacataaaatttc 1002
SEQIDNO4 cctcaacccctttttctccaacaaaagaatgcaaacagaacttgaataacataaaatttc 1020
************************************************************
TFL1bpromoter acatgcagagatgaagagggcaaaatggtttcttgataaagaaggtccaccttcctcttt 1062
SEQIDNO4 acatgcagagatgaagagggcaaaatggtttcttgataaagaaggtccaccttcctcttt 1080
************************************************************
TFL1bpromoter taccaatattggaacttggtctgcatctaagagttaagtttttattctgaaatgacctct 1122
SEQIDNO4 taccaatattggaacttggtctgcatctaagagttaagtttttattctgaaatgacctct 1140
************************************************************
TFL1bpromoter taggcactcaaacgcttatactccataggcgtgtctagccttgagaatttgtggtggagt 1182
SEQIDNO4 taggcactcaaacgcttatactccataggcgtgtctagccttgagaatttgtggtggagt 1200
************************************************************
TFL1bpromoter tcctttaattatgccctcttcatcaggaatcttccaacaaaattaaaagaaagactatgg 1242
SEQIDNO4 tcctttaattatgccctcttcatcaggaatcttccaacaaaattaaaagaaagactatgg 1260
************************************************************
TFL1bpromoter ctaagggaaaatggagaatcacgcacgcacaaacagaaatttgcatagtgtgtgcaaaca 1302
SEQIDNO4 ctaagggaaaatggagaatcacgcacgcacaaacagaaatttgcatagtgtgtgcaaaca 1320
************************************************************
TFL1bpromoter attaaggaaggttctctacgcaaggcatacaacagaaatttctgcacgtccatggtacta 1362
SEQIDNO4 attaaggaaggttctctacgcaaggcatacaacagaaatttctgcacgtccatggtacta 1380
************************************************************
TFL1bpromoter gtagtatactaaagagaatgctaaaatgtgggtgttcaagaaacgaagcatttcctagaa 1422
SEQIDNO4 gtagtatactaaagagaatgctaaaatgtgggtgttcaagaaacgaagcatttcctagaa 1440
************************************************************
TFL1bpromoter tttttgccttttatgcttttagtttaataataaaacggcagaagcttgacaacttgcaaa 1482
SEQIDNO4 tttttgccttttatgcttttagtttaataataaaacggcagaagcttgacaacttgcaaa 1500
************************************************************
TFL1bpromoter ggggagggaggacccacacacacaggattacacatgatgctttttttgacctgatcgagc 1542
SEQIDNO4 ggggagggaggacccacacacacaggattacacatgatgctttttttgacctgatcgagc 1560
************************************************************
TFL1bpromoter tgatgggagagaaaaaactgggaaattttatatttttaaatattttttacaaattttttc 1602
SEQIDNO4 tgatgggagagaaaaaactgggaaattttatatttttaaatattttttacaaattttttc 1620
************************************************************
TFL1bpromoter agaactaggatgggatagaaataaataaaaaaagtacacaataagagttgctaatagaat 1662
SEQIDNO4 agaactaggatgggatagaaataaataaaaaaagtacacaataagagttgctaatagaat 1680
************************************************************
TFL1bpromoter aaaaaaatttctttattggatgagaaatatccatgcatccggtgtacagccaatcaatta 1722
SEQIDNO4 aaaaaaaattctttattggatgagaaatatccatgcatccggtgtacagccaatcaatta 1740
******* ****************************************************
TFL1bpromoter agaaccgattccaagggccccattctgtgtttatgagcagcggagagagaaagagagtgg 1782
SEQIDNO4 agaaccgattccaagggccccattctgtgtttatgaccagcggagagagaaagagagtgg 1800
************************************ ***********************
TFL1bpromoter aaaagtggaaaggataactcgtcacatatcatgactaatgtgatttcagcacaaacatcc 1842
SEQIDNO4 aaaagtggaaaggataactcgtcacatatcatgactaatgtgatttcagcacaaacatcc 1860
************************************************************
TFL1bpromoter cagacccaaaggcaattgttgagatgagccgaattaagagacaatatattagtagtagtg 1902
SEQIDNO4 cagacccaaaggcaattgttgagatgagccgaattaagagacaatatattagtagtagtg 1920
************************************************************
TFL1bpromoter ggaggacctgcacggatgcacttgatttatttctagccttttttcattttttgtttttcc 1962
SEQIDNO4 ggaggacctgcacggatgcacttgatttatttctagccttttttcattttttgtttttcc 1980
************************************************************
TFL1bpromoter tatagaacttgtattttctctagctaaatctaaattatgtattgcaacatattttaaagg 2022
SEQIDNO4 tatagaacttgtattttctc---------------------------------------- 2000
********************
TFL1bpromoter aggggcaaaacacactcgatctaccttcaaagaaacttttaagacattataaatatggag 2082
SEQIDNO4 ------------------------------------------------------------ 2000
TFL1bpromoter cccacactcccttcttcctcagtaagatcatcaacttgtattcctcacttccttagctcc 2142
SEQIDNO4 ------------------------------------------------------------ 2000
TFL1bpromoter tcctcttccttgttcctcctcttaca 2168
SEQIDNO4 -------------------------- 2000
Scope of Enablement
Claims 12-15 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 a limited set of promoter deletions in a specific soybean Ct1/GmTFL1b promoter corresponding to the disclosed 2 kb promoter sequence (SEQ ID NO:4), and confirming the edits and changes in GmTFL1b expression by molecular assays; does not reasonably provide enablement for obtaining across the full scope of claimed promoter edits plants that consistently meet the additional agronomic limitations of claims under diverse genetic backgrounds and environmental/pathogen conditions. 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.
Claims 12-15 regards additional agronomic performance limitations, namely, earlier terminal flowering, reduced lodging, substantially the same or increased yield, and lower fungal disease susceptibility, relative to an otherwise identical control.
Breath of the claims
The claims encompass a broad set of distinct promoter modifications (multiple deletion sizes/endpoints and combinations) and apply the outcome limitations of claims 12-15 to the entire breadth of those edits (Claim 1 Markush list and “combinations”, p0136 table 2). And the underlying “TFL1b gene” scope is further expanded by claim 2’s broad sequence-identity definition. This breadth encompasses a large number of distinct promoter alleles and genetic contexts, such that identifying which embodiments (if any) reliably produce earlier terminal flowering, reduced lodging, maintained/increased yield, reduced fungal disease susceptibility would require extensive screening and testing beyond routine experimentation. Accordingly, the claims are broad relative to the experimental disclosure.
Unpredictability/nature of the invention
The recited limitations of claims 12-15 are complex agronomic traits that are influenced by multiple factors, including genetic background and environmental conditions, and therefore are not predictable solely from the presence of a promoter deletion or from expression measured as a single developmental stage. For example, lodging and yield depend on multiple interacting plant architecture and environmental viables, and fungal disease susceptibility further depends on pathogen presence/pressure and microclimate conditions (instant application, p0150). Furthermore, the specifications’ discussion of fungal disease susceptibility appears to be speculative and indirect, attributing any potential reduction in fungal infections to secondary effects such as altered plant architecture (e.g., a more compact, upright canopy improving airflow and reducing humidity) rather than to any demonstrated or mechanistically support role of the endogenous TFL1b/Dt1 gene or its promoter in fungal defense pathway (instant application p0150).
Amount of direction/presence of working examples
The specification primarily describes these claim 12-15 outcomes in prophetic or expected terms (e. g., plant “will be evaluated” (p0151, 0142, 0143); “it is expected” (p0145, 0158); “has the potential”(p0145); “potential for decreased fungal disease susceptibility” (p0150) ), and does not provide sufficient experimental data demonstrating that the claimed edits, across their breadth, produce the required phenotypes (earlier terminal flowering, reduced lodging, maintained/increased yield, reduced fungal disease susceptibility).
In view of the breadth of the promoter edits encompassed by the claims, the unpredictability of the recited agronomic traits, and the limited guidance and lack of working examples commensurate with the full scope, a person of ordinary skill would be required to perform undue experimentation, including extensive multi-environment field testing and trait screening across numerous edits and genetic backgrounds, to identify embodiments that satisfy claims 12-15
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 1-17 are rejected under 35 U.S.C. §103 as being unpatentable over Liu (2010) (Baohui Liu et. al., 2010 May;153(1):198-210, 2010 Mar 10), in view of Liu (2016) (Yunfeng Liu et. al., Innovation of a Regulatory Mechanism Modulating Semi-determinate Stem Growth through Artificial Selection in Soybean, PLOS Genetics DOI:10.1371/ journal.pgen.1005818 January 25, 2016), and further in view of Duan (Kaixuan Duan et. al., Large chromosomal segment deletions by CRISPR/LbCpf1‐mediated multiplex gene editing in soybean, Journal of Integrative Plant Biology, September 2021 | Volume 63 | Issue 9 | 1620–1631).
Claim 1 recites a modified soybean plant/seed/part/cell comprising a modification in a promoter of an endogenous TFL1b gene, wherein the modification reduces expression or activity of TFL1b relative to an otherwise identical control lacking the modification, and wherein the modification is selected from a Markush group of specified promoter deletions (and combinations).
As discusses above, since the limitation of “endogenous TFL1b gene” is unclear, claim 1 is interpreted with the scope of anything that is called TF1b.
Liu(2010) teaches that the soybean Dt1 locus encodes the TERMINAL FLOWER 1 ortholog GmTFL1b (TFL1b) and that Dt1/TLF1b is a key genetic determinant of soybean stem growth habit/determinacy (i. e. , alteration/reduction of Dt1/TFL1b function is associated with increased determinacy) (p198, abstract and introduction).
Liu(2016) teaches that Dt1 (GmTFL1b/TFL1 ortholog) determinacy is controlled by transcriptional regulation via its promoter(e. g., promoter control by transcription factors). (p1-2, abstract and introduction); identifies and uses an ~2.4 kb upstream Dt1 promoter region (p16); and demonstrates that changing that promoter sequence (truncation/deletions or small base changes in regulatory motifs) modulates Dt1 promoter activity and Dt1 expression (p16, p6, fig 2). Liu identifies CArG boxes in ~2 kb upstream promoter region of Dt1, and shows 1st, 2nd, 5th CArG boxes binds to Dt2 proteins (fig 2); Liu makes both promoter truncation/deletion removing CArG boxes, and small point mutations (3-4nt) in individual CArG boxes to determine promoter function (fig 5, fig 7). In conclusion, Liu tests TF-binding +cis-element mutation /truncation assays demonstrating Dt1 expression can be repressed in semi-determinate soybean.
Liu(2010) and Liu(2016) do not teach introducing the claimed specific promoter deletions (as recited by the nucleotide coordinates relative to SEQ ID NO:4) in soybean using genome editing.
A POSITA would have been motivated to modify the Dt1/TFL1b promoter to modulate (including reduce) Dt1/TFL1b expression to obtain determinacy-related phenotypes.
Duan teaches that CRISPR/LbCpf1 (Cas12a/Cpf1) multiplex editing in soybean using multiple guide RNAs can generate targeted deletions, including small deletions at cut sites and segment deletions between two cut sites, and that such deletion alleles are routinely identified by screening (p1621, abstract).
Therefore, it would have been obvious to one of ordinary skill in the art to modify the promoter of endogenous Dt1/TLF1b in soybean (as motivated by Liu(2010)/Liu(2016)) using the Cpf1-based deletion strategy of Duan, with a reasonable expectation of success, to obtain promoter deletion alleles that reduce Dt1/TLF1b expression/activity relative to a corresponding control lacking the modification, as required by claim 1. Because CRISPR/Cpf1 editing inherently yields a spectrum of deletion endpoints around the cut sites, and because the position/extent of promoter deletions is a result-effective variable for modulating Dt1/TLF1b transcription, it would have been obvious to generate and routinely screen multiple promoter deletions within the targeted Dt1 promoter region and select particular deletion sizes/endpoints.
Claim 2 recites the TFL1b gene encodes a protein having a listed range of sequence identity to SEQ ID NO:2.
Liu(2010) teaches the Dt1-encoded GmTFL1b protein sequence is 97% homologous to SEQ ID NO:2. GmTFL1b sequence can be found from GenBank/EMBL/DDBJ data libraries under accession numbers AB511820 (page 208, above supplemental data). (see sequence alignment below). Therefore, claim 2 would have been obvious for the same reasons as claim 1.
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Claim 3 depends from claim 1 and recites the plant, plant seed, plant part, or plant cell is heterozygous for the modification.
It is routine in plant genetics and genome-editing workflows (including soybean) to recover and identify heterozygous and homozygous edited genotypes through selfing and selection after transformation/editing. It would have been obvious that a genome-edited soybean initially recovered after introducing a targeted promoter modification would commonly be heterozygous for the edit, because targeted nuclease editing is typically introduced into a diploid genome and edits are frequently obtained on only one allele in early regenerated events, determining whether and edited plant is heterozygous is accomplished by routine genotyping/sequencing. Claim 3 is obvious over Liu(2010), in view of Liu(2016) and Duan.
Claim 4 depends from claim 1 and recites the plant, plant seed, plant part, or plant cell is homozygous for the modification.
Claim 4 is obvious for the same reasons set forth with respect to claim 1. It would have been obvious to obtain a homozygous promoter-edited soybean from an edited event by selfing and selecting progeny with desired genotype, since producing homozygous lines from a heterozygous edited plant is a routine and predictable step in plant genetics and breeding, and the desired genotype is readily identified by routine molecular assays.
Claim 5 depends from claim 1 and recites the plant, or plant part comprises a first modification in a first allele of the TFL1b gene and a second modification in a second allele of the TFL1b gene, the first modification and the second modification being different from one another.
It would have been obvious that the two alleles of a targeted locus can carry different edits following targeted nuclease-induced double-strand breaks, because repair outcomes can occur independently on each allele, producing distinct deletion endpoints, identifying plants having different edits on different alleles is a routine matter of sequencing/genotyping and selection.
Claim 6 depends from claim 1 and recites wherein the plant, plant seed, plant part, or plant cell comprises a chromosomal sequence in the TFL1b gene that has at least 70% to 100% sequence identity to SEQ ID NO:4 in the regions outside of the modification.
Because in Dt1/TFL1b promoter-edited soybean of claim 1, the modification is limited to the recited promoter deletions, and therefore the chromosomal sequence outside the modification would inherently remain substantially identical to the corresponding reference promoter sequence (SEQ ID NO:4), satisfying the claimed sequence-identity requirement. Claim 6 is obvious in view of Liu(2010), Liu(2016) and Duan.
Claim 7 depends from claim 1 and recites wherein the plant, plant seed, plant part, or plant cell comprises a polynucleotide sequence selected from the group consisting of SEQ ID NOs: 12- 39.
SEQ ID NOs: 12-39 are polynucleotide sequences for alleles of the GmTFL1b promoter having various deletions as compared to SEQ ID NO:4. Because once the Dt1/TFL1b promoter is targeted for genome editing to modulate Dt1/TFL1b expression, the Cpf1-based editing approach predictably yields a spectrum of promoter deletion alleles that are identified by routine sequencing/genotyping, and selecting particular edited promoter sequences (including those corresponding to SEQ ID NOs: 12-39) is an obvious results of routine screening and optimization of deletion endpoints within the targeted promoter region.
Claim 8 depends from claim 1 and recites wherein the modification alters the expression level of the TFL1b protein, as compared to the expression level of the TFL1b protein in an otherwise identical plant, plant seed, plant part, or plant cell that lacks the modification.
Claim 8 is obvious for the same reasons set forth with respect to claim 1. Because modifying the Dt1/TFL1b promoter as motivated by Liu(2010) and Liu(2016) using the genome-editing approach of Duan, would have been expected to alter the expression level of the TFL1b protein relative to an otherwise identical plant lacking the promoter modification.
Claim 9 depends from claim 8 and recites the modification reduces the expression level of the TFL1b protein, as compared to the expression level of the TFL1b protein in an otherwise identical plant, plant seed, plant part, or plant cell that lacks the modification.
Because Liu(2010) establishes Dt1/TFL1b as determinacy gene and Liu(2016) indicates Dt1 is regulated through its promoter such that promoter changes can decrease Dt1/TFL1b expression; applying the Cpf1-based promoter deletion approach in soybean to the Dt1/TFL1b promoter would have made it obvious to obtain and select prompter-edited plants in which the modification reduces the expression level of TFL1b relative to an otherwise identical plant lacking the modification.
Claim 10 depends from claim 1 and recites the modification alters the determinacy phenotype of the plant, as compared to the determinacy phenotype of an otherwise identical plant that lacks the modification.
Because Liu(2010) teaches that Dt1/TFL1b controls soybean stem growth habit/determinacy (p198) and Liu(2016) teaches Ct1 is regulated through it promoter such that altering the promoter modulates Dt1/TFL1b expression and associated growth-habit phenotypes (fig 6, 7); therefore, generating Dt1/TFL1b promoter edits using the Cpf1-based deletion approach of Duan would have been expected to alter the determinacy phenotype of the plant relative to an otherwise identical plant lacking the modification.
Claim 11 depends from claim 10 and recites modification increases the determinacy of the plant, as compared to the determinacy of an otherwise identical plant that lacks the modification,
Because Liu(2010) teaches that reduced Dt1/TFL1b function is associated with increased determinacy and Liu(2016) teaches Dt1 expression is controlled by its promoter such that promoter modifications can reduce Dt1/TFL1b expression, therefore, introducing promoter deletions in the Dt1/TFL1b promoter using the Cpf1-based approach of Duan and selecting the alleles that decrease expression would have been expected to increase the determinacy of the plant relative to an otherwise identical plant lacking the modification.
Claim 12 depends from claim 1 and recites after planting, the modified plant reaches its terminal flowering date sooner than an otherwise identical plant that lacks the modification.
Because Liu(2010) teaches Dt1/TFL1b regulates termination/determinacy and Liu(2016) teaches Dt1 expression is controlled by its promoter such that decreasing Dt1/TFL1b expression increases determinacy, therefore, generating Dt1/TFL1b promoter deletions using the Cpf1-based method of Duan and selecting edits that reduce expression would have been expected to cause the modified plant to reach terminal flowering sooner than an otherwise identical plant lacking the modification.
Claim 13 depends from claim 1 and recites the modified plant exhibits a reduced lodging rate, as compared to the lodging rate of an otherwise identical plant that lacks the modification.
Because Liu(2010) and Liu(2016) teach that reducing Dt1/TFL1b expression alters soybean stem growth habit toward increased determinacy, which would have been expected to produce a more compact/less indeterminate plant architecture, and it would have been obvious to select among the promoter-edited plants produced by the Cpf1-based editing approach of Duan those exhibiting improved standability, i. e., a reduced lodging rate, relate to an otherwise identical plant lacking the modification.
Claim 14 depends from claim 1 and recites the modified plant exhibits substantially the same or increased yield, as compared to the yield of an otherwise identical plant that lacks the modification.
Because Liu(2010) and Liu(2016) teach modifying Dt1/TFL1b expression via promoter regulation to alter determinacy/growth habit, and it would have been obvious to one of ordinary skill in the art to generate a population of Dt1/TFL1b promoter-edited soybean plants using the Cpf1-based approach of Duan and to select promoter-edit alleles that achieve the desired determinacy phenotype while maintaining yield, such that the modified plant exhibits substantially the same or increased yield compared to an otherwise identical plant lacking the modification.
Claim 15 depends from claim 1 and recites the modified plant exhibits lower susceptibility to fungal disease, as compared to the susceptibility of an otherwise identical plant that lacks the modification.
Because Liu(2010) and Liu(2016) teach that altering Dt1/TFL1b expression changes soybean growth habit and plant architecture, and it would have been obvious to generate Dt1/TFL1b promoter-edited soybean plants using the Cpf1-based editing approach of Duan and to select lines with advantageous agronomic traits associated with altered architecture, including lower susceptibility to fungal disease, relative to an otherwise identical plant lacking the modification.
Claim 16 recites a polynucleotide comprising a sequence selected from the group consisting of SEQ ID NOs: 12-39.
SEQ ID NOs: 12-39 are polynucleotide sequences for alleles of the GmTFL1b promoter having various deletions as compared to SEQ ID NO:4. Because once the Dt1/TFL1b promoter is targeted for genome editing to modulate Dt1/TFL1b expression, the Cpf1-based editing approach predictably yields a spectrum of promoter deletion alleles that are identified by routine sequencing/genotyping, and selecting particular edited promoter sequences (including those corresponding to SEQ ID NOs: 12-39) is an obvious results of routine screening and optimization of deletion endpoints within the targeted promoter region.
Claim 17 recites a guide RNA comprising a polynucleotide sequence selected from the group consisting of SEQ ID NOs: 10 and 11.
Because once Dt1/TFL1b promoter is selected as a target to modulate determinacy-related expression (Liu2010, Liu2016), and Cpf1-based genome editing in soybean using guide RNAs is applied (Duan, p1626), designing guide RNAs that target selected positions within the Dt1/TFL1b promoter is a routine and predictable implementation details (selection of protospacers adjacent to a suitable PAM and minimizing off-targets), and therefore, using guide RBAs corresponding to SEQ ID Nos:10 and 11 would have been an obvious choice for carrying out the promoter-editing strategy.
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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/YANXIN SHEN/ Examiner, Art Unit 1663
/WEIHUA FAN/ Primary Examiner, Art Unit 1663