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 .
Information Disclosure Statement
The Information Disclosure Statements filed on 7/16/2025 have been entered and considered. Initialed copies of the form PTO-1449 are enclosed with this action.
Status of claims
Applicant’s response filed 9/15/2025 has been entered.
Note: On 3/3/2025, the applicant elected Group I, claims 1-9, also elected species of Election 1--NK603, Election 2--DP-4114. MIR162 was rejoined in Election 1; MIR604 was rejoined in Election 2.
Claims 8-9 have been canceled by the applicant.
Claims 1, 6-7 have been amended.
In summary, claims 1-7 are examined in this office action. Non-elected claims and species are withdrawn.
All previous objections and rejections not set forth below have been withdrawn in view of the applicant’s amendment and/or upon further consideration. See “Response to Arguments” at the end of office action. The following rejections are repeated, modified and/or added for the reasons of record as set forth in the last Office action of 5/16/2025, and/or necessitated by the applicant’s amendments. The applicant’s arguments filed 9/16/2025 have been thoroughly considered but are not deemed fully persuasive.
Claim Objections
The amended claim 1 is objected for the informality:
The claim improperly uses “or” in the Markush group.
In the 3rd from the last line, it is suggested to change “or a modification there of ……” to --- and a modification there of …… ---.
Appropriate correction is required.
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries set forth in Graham v. John Deere Co., 383 U.S. 1, 148 USPQ 459 (1966), that are applied for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or non-obviousness.
Claims 1-7 are rejected under 35 U.S.C. 103 as being unpatentable over Long et al (US 8232456, granted and published 7/31/2012), in view of Du et al (Construction of Marker-Free Genetically Modified Maize Using a Heat-Inducible Auto-Excision Vector. Genes, p1-17, 2019), Cai et al (CRISPR/Cas9-Mediated Deletion of Large Genomic Fragments in Soybean. Int. J. Mol. Sci. 19, 1-13, 2018), Qi et al (High-efficiency CRISPR/Cas9 multiplex gene editing using the glycine tRNA-processing system-based strategy in maize. BMC Biotechnology, p1-8, 2016), and references teaching the particular elected species (MIR162 OR MIR604 OR NK603 OR DP-4114): EFSA GMO Panel 2019 (Assessment of genetically modified maize Bt11 x MIR162 x MIR604 x 1507 x 5307 x GA21 and subcombinations, for food and feed uses, under Regulation (EC) No 1829/2003 (application EFSA-GMO-DE-2011-103). EFSA Journal, p1-35, 2019), EFSA GMO Panel 2017 (Assessment of genetically modified maize 1507 x 59122 x MON810 x NK603 and subcombinations, for food and feed uses, under Regulation (EC) No 1829/2003 (application EFSA-GMO-NL-2011-92). EFSA Journal, p1-29, 2017), and Diehn et al (US Patent 8575434, granted and published 11/5/2013).
Amended claim 1 is drawn to a method comprising steps of:
(i) inducing at least one targeted genetic change in the genome of the elitemaize plant with one or more genome editing molecules, in an the genome of the elite maize plant comprising a first approved transgenic locus or modification thereof comprising a deletion of at least one selectable marker gene and/or non-essential DNA, and a second approved transgenic locus or modification thereof comprising a deletion of at least one selectable marker gene and/or non-essential DNA wherein the at least one targeted genetic change is induced at a location in the genome of the elite maize plant other than the first approved transgenic locus;
(ii) excising a DNA segment comprising all of the first approved transgenic locus with genome editing molecules by contacting genomic DNA of said plant with:
(a)at least a first and at least a second guide RNA directed to genomic DNA adjacent to two protospacer adjacent motif (PAM) sites, wherein the PAM sites are operably linked to a 5' and a 3' DNA junction polynucleotide of the first approved transgenic locus; and
(b) one or more RNA dependent DNA endonucleases (RdDe) which recognize the PAM sites; and
(iii) selecting an elite maize plant wherein the first approved transgenic locus or modification thereof is excised, the second approved transgenic locus or modification thereof is present, and the targeted genetic change is present;
wherein the first approved transgenic locus and the second approved transgenic locus are not the same and are independently selected from MIR162, MIR604, NK603, or DP-4114 (elected species), or a modification thereof comprising a deletion of at least one selectable marker gene and/or non-essential DNA,
for producing an elite maize plant comprising a targeted genetic change and at least one approved transgenic locus (preamble).
Interpretation of claim 1
According to dictionaries, a or each locus refers to the physical location of a specific gene on a chromosome.
In another word, if multiple genes are located in a region of a chromosome, such region comprises multiple loci, and is not a single locus.
In view of prior art, MIR162 (long et al above), NK603 (US Patent 6825400, granted and published 11/30/2004), and DP-4114 (Pioneer_2014), all are elite transgenic events, each of these events contains multiple transgenic loci, and each locus comprises a transgenic. Thus, the elected MIR162, NK603, MIR604 or DP-4114R is not a locus: each of them is a transgenic event comprising multiple loci. See the rejection below. Removing a whole elite transgenic event would not have any scientific sense or any commercial benefit.
In addition, in view of the specification (Example 1 in [00244]-[00249], Tables 5-6; Example 1 is the only example in the specification that is not a working example), the applicant intends to remove marker genes (PAT, EPSPS, PMI, and HPT) from within the transgenic events MIR162, NK603, DP-4114 (in Tables 5-6, they are called “MAIZE EVENT NAME”, not locus name) and so on, not the whole multi-loci transgenic event.
Furthermore, claim 1 requires (iii) selecting an elite maize plant wherein the first approved transgenic locus (marker gene and 5’ and 3’ junctions as recited in claim 1) is excised, the second approved transgenic locus (resistant gene or other gene of interest also as recited in claim 1) is present, and the targeted genetic change is present.
Thus, for compact prosecution, by BRI and from the context of the claim, the last wherein clause of claim 1 is interpreted as:
wherein the first approved transgenic locus and the second approved transgenic locus are not the same and are independently selected from within DP-4114 event, MIR162 event, MIR604 event, or NK603 event, or a modification thereof comprising a deletion of at least one selectable marker gene and/or non-essential DNA.
Note: Each of DP-4114, MIR162, MIR604, NK603, or a modification thereof, is an option, not requires. Each comprises multiple and different genes in multiple and different loci (gene of interest and marker gene for example).
Long et al teach making a transgenic maize event designated MIR162 (abstract), and the plant comprising MIR162 event (abstract; col 3, last para; claims 1-2). Long deposited the seed comprising MIR162 (ATCC PTA-8166) (Example 10 in col 40). The transgenic maize of Long et al is an elite maize plant.
Long et al teach that the transgenic maize plant cell comprises a nucleotide sequence comprising a phosphomannose isomerase (PMI, a selectable marker) coding region and the own driving promoter (reading on first transgenic locus comprising a selectable marker gene); and a vip3Aa19 (insect resistance) coding region and the own driving promoter (reading on second approved transgenic locus) (Example 1 in col 21, lines 40-67, and col 22, lines 1-20). Both PMI gene and vip3Aa19 coding gene are well known in the art, thus, each of them is an approved transgenic locus.
Long et al also teach inducing at least one targeted genetic change (like substitution) in the genome of the maize plant with one or more genome editing molecules in one of the loci. The changed part is in Vip3A locus of the event of the maize genome. Thus, the changed part is in the second, not the first, locus (col 17, lines 28-67).
The locus (locus 1) comprising PMI gene and the locus (locus 2) comprising the Vip3A gene are not the same, and are independently selected, and are within MIR162 (one of the options and one of the elected species).
Thus, Long et teach step (i) including the new limitation that the targeted genetic change is induced at a location other than the first approved transgenic locus, and including that the first transgenic locus (comprising PMI marker gene) and the second transgenic locus (comprising Vip3A gene of interest) are not the same and are within the MIR162 event (an event comprising multiple and different loci, not a locus, one of the elected species).
Long et al do not steps (ii) and (iii).
Du et al teach and demonstrated a detailed method of site-specific removal/excising a marker gene completely from transgenic maize by using excision vector (p1, abstract; p3, fig 1; Materials and Methods in p4-9, whole pages). Du et al teach removing sugar metabolism related PMI gene (p2, 1st para; PMI gene is the marker gene of Long et al).
Du et al teach and demonstrated success in selecting resultant maize plant that comprises marker-free transgenic locus, which reads on an elite maize plant wherein the first approved transgenic locus is excised, the second approved transgenic locus is present, and the targeted genetic change is present (Results in p8-11).
Du et al further teach that production of marker-free transgenic crops is crucial to avoid potential risks and promote commercial deployment of GM plants by hastening public acceptance (p2, 2nd para), the advantage of excising the marker gene.
Thus, Long et al in view of Du et al teach steps (i)-(iii) and achieved success of the preamble, except
do not teach part of the step (ii) that the excising is by contacting genomic DNA of said plant with: (a) at least a first and at least a second guide RNA directed to genomic DNA adjacent to two protospacer adjacent motif (PAM) sites, wherein the PAM sites are operably linked to a 5' and a 3' DNA junction polynucleotide of the first approved transgenic locus; and (b) one or more RNA dependent DNA endonucleases (RdDe) which recognize the PAM sites.
Cai et al teach deleting/excising marker gene (the first approved transgenic locus) with genome editing molecule (CRISPR-Cas9), and guide RNA directed to genomic DNA adjacent to two protospacer adjacent motif (PAM) sites, operably linked to a 5' and a 3' DNA junction of the DNA sequences comprising a marker gene (the location of the marker gene reads on the first approved transgenic locus) (p1, abstract; p3, figure 1, p5, figure 2).
Cai et al teach that deletions of regulatory elements/sequences (5' and/or 3' DNA junction of the DNA sequences comprising a gene) increase the effectiveness of gene editing (p9, 2nd para; p10, 2nd para), and that the PAM sites are particularly effective for deletion of large sequences 599 bp to 4.4 kb leading to knockout of genes (p9, 2nd para), an advantage of PAM sites. Cai et al teach locating PAM sequences in plants (p5, figure 2).
As a support of Cai et al, Qi et al teach locating PAM sequences in maize (p5, fig 3). Qi et al teach deleting/excising DNAs/genes with CRISPR-Cas9 and guide RNA directed to genomic DNA adjacent to two protospacer adjacent motif (PAM) sites, operably linked to a 5' and a 3' DNA junction of the DNA sequences, and achieved success in maize (Methods and Results in p2-6, figs 1-4). Qi et al teach that the particular CRISPR-Cas9 and guide RNA system contains one or more RNA dependent DNA endonucleases (RdDe) for recognition of the PAM sites (p1, left col, 1st para; p7, left col, 1st para).
Regarding dependent claims, Du et al teach that the removing/excising the marker gene is performed after transformation/genetic change (page 2, 2nd para), teaching the limitation of claim 2.
Cai et al teach that the targeted genetic change and gene deletion can be simultaneously achieved (page 2, 1st para), teaching the limitation of claim 3.
Long et al teach that the MIR162 event comprising the vip3Aa19 gene of Long confers insect resistance, a desirable agronomic trait (Abstract; col 3, lines 38-67), teaching the limitation of claim 4.
Regarding dependent claims 5-6, Cai et al teach that the deletion/excision between PAM sites including the deletion of the genes and deletion/excision of the 5' and 3' DNA junction polynucleotides of the genes. The deletion of the junction polynucleotides is not 100% (reads on all but at least one nucleotide of the heterologous DNA of the 5' and/or 3' DNA junction polynucleotide of the first approved transgenic locus is excised) (page 5, figure 2).
Long et al teach obtain obtaining the elite crop plant comprising a first approved transgenic loci (PAM gene locus) and a second approved transgenic locus (Vip3Aa19 gene locus) of step (i), teaching the limitation of claim 7.
An invention would have been obvious to one ordinary skill in the art if any teaching, suggestion or motivation in prior art leading the one to combine the teaching(s) or suggestion(s) of the cited references to arrive the claimed invention.
It would have been obvious for one ordinary skill in the art to modify the invention of Long et al such that the PMI marker gene is excised from the elite maize of Long et al, and the gene of interest (Vip3A gene) is present, as taught by Du et al. One having ordinary skill in the art would have been motivated to do so because removing selectable marker genes is crucial for commercial genetically modified maize development as taught Du et al.
It also would have been obvious for one ordinary skill in the art to use the methods as taught collectively by Cai et al and Qi et al to perform such excision of Du et al such that the PAM sites are used, and the 5’ and 3’ junction sequences of the marker gene are also removed. One having ordinary skill in the art would have been motivated to do so increases effectiveness of the editing.
The expectation of success would have been high, because excision of marker genes had been a taught and demonstrated methods, for example by Du et al and Cai et al. By finding and using PAM sites and PAM recognizing sequences as taught and demonstrated by Cai et al and Qi et al collectively, the expectation of success would have been enhanced.
Therefore, the invention would have been obvious to one ordinary skill in the art.
Remarks
For compact prosecution, if the first transgenic event and the second transgenic event were not the same and independently selected, Long et al further suggest that maize line comprising MIR162 event can cross with another elite maize line. Specifically, maize line comprising MIR162 event can cross with elite maize line comprising MIR604 event, such that the progeny maize line comprises both MIR162 and MIR604 events (col 16, lines 47-67; col 17, lines 1-10). Long et al teach that MIR604 event comprises gene encoding insect resistance cry3A055 (col 36, 28-40).
EFSA GMO Panel 2019 (Assessment of genetically modified maize Bt11 x MIR162 x MIR604 x 1507 x 5307 x GA21 and subcombinations, for food and feed uses, under Regulation (EC) No 1829/2003 (application EFSA-GMO-DE-2011-103). EFSA Journal, p1-35, 2019, the PDF was attached in the non-final rejection of 5/16/2025) explicitly teaches and demonstrated making a new maize line comprising multiple transgenic events including both MIR162 and MIR604 events (p1, Abstract). EFSA GMO Panel concludes that the environmental impact of the new maize line is likely low (p29, 3rd para).
The MIR604 event, like MIR162 event, also comprises multiple and different loci, some locus comprises gene of interest, some different locus comprises marker gene (p11-12, Table 4).
Thus, the subject matter would still be deemed obvious.
The following references are relevant to instant application:
EFSA GMO Panel 2017 (Assessment of genetically modified maize 1507 x 59122 x MON810 x NK603 and subcombinations, for food and feed uses, under Regulation (EC) No 1829/2003 (application EFSA-GMO-NL-2011-92). EFSA Journal, p1-29, 2017).
Diehn et al (US Patent 8575434, granted and published 11/5/2013).
Behr et al (US Patent 6825400, granted and published 11/30/2004).
The PDFs were attached in the non-final rejection of 5/16/2025.
Chen et al (CA 3069542, published 7/26/2018, filed 2/7/2029). PDF attached.
Response to Arguments
In view of the significant amendment to claim 1, the 103 rejection is re-written. Thus, the arguments to the previous rejection are not longer applicable.
The examiner would emphasize again (as did in the previous office action) that a transgenic event like MIR162 is not a locus. It comprises multiple and different loci instead. For example, MIR162 comprises a locus locating a PMI gene, and comprises another locus locating Vip3A gene. The locus locating a PMI gene, and the locus locating Vip3A gene, are two different loci.
Conclusion
No claim is allowed.
The applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). The applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any extension fee pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the date of this final action.
Contact information
Any inquiry concerning this communication or earlier communications from the examiner should be directed to WAYNE ZHONG whose telephone number is (571)270-0311. The examiner can normally be reached 8:30am to 5:00pm EST.
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/Wayne Zhong/
Primary Examiner, Art Unit 1662