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 9/2/2026 have been entered and considered. Initialed copies of the form PTO-1449 are enclosed with this action.
Status of claims
Applicant’s response filed 8/26/2026 has been entered.
Claims 2, 5-11, 14-16, 19-22, 24, 27-38, 51-59, 63-64, 66-67 had/have been canceled by the applicant.
Claims 1, 3-4, 23, 62, 65 have been amended.
In summary, claims 1, 3-4, 12-13, 17-18, 23, 25-26, 39-40, 47-50, 60-62, 65 are pending and examined in the office action.
All previous objections and rejections not set forth below have been withdrawn in view of the applicant’s amendment and/or upon further consideration.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 7/22/2026, and/or necessitated by the applicant’s amendments. The applicant’s arguments filed 8/26/2026 have been thoroughly considered but are not deemed fully persuasive.
Definition of "dry-excised mature plant embryo explant" in the Specification
In the specification ([0022]), "A "dry excised explant" is a mature embryo explant taken or excised from a mature dry seed. A plant seed naturally dries during its maturation process. As described further below, other types of explants may be taken or excised from a mature seed depending on their treatment, such as after wetting, imbibing, etc., a dry seed, and an explant may be wetted, imbibed, etc., after being excised from a dry seed”.
Accordingly, after being wetted, imbibed, etc., such explant still reads on a "dry-excised mature plant embryo explant". Otherwise, the claims may have an enablement issue: bacterially mediated transformation requires co-culturing of the bacteria and the seeds or explants in a medium/solution.
In addition, the specific also does not provide any example of “delivering to a dry-excised mature plant embryo explant via bacterially mediated transformation a recombinant DNA construct”. Examples 1-5, 7 are examples of particle bombardment. Example 6 is an example of identification.
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, 3-4, 12-13, 17-18, 23, 25-26, 39-40, 47-50, 60-61 are rejected under 35 U.S.C. 103 as being unpatentable over Shultz et al (WO 2018085693, published 5/11/2018, filed 11/3/2017, priority 11/4/2016 to US62418078), in view of Patnaik et al (Agrobacterium-mediated transformation of mature embryos of Triticum aestivum and Triticum durum. Current Science. 91, 307-317, 2006), Li et al (Optimization of Agrobacterium-Mediated Transformation in Soybean. Frontiers in Plant Science. P1-15, 2017), and Char et al (An Agrobacterium-delivered CRISPR/Cas9 system for high-frequency targeted mutagenesis in maize. Plant Biotechnology Journal. 15, pp. 257–268, 2017).
Amended claim 1 is drawn to a method comprising:
a) excising an embryo from a dry mature plant seed, thereby producing a dry-excised mature plant embryo explant;
b) delivering to the dry-excised mature plant embryo explant via bacterially mediated transformation a recombinant DNA construct comprising a sequence encoding a site- specific nuclease, wherein the sequence is operably linked to a plant-expressible promoter; and
c) regenerating a plant from the dry-excised mature plant embryo explant, wherein the regenerated plant comprises an edit or site-directed integration at or near the target site of the site-specific nuclease in the genome of at least one cell of the regenerated plant.
for editing a genome of a plant (preamble).
Claim 3 limits the transformation to Agrobacterium mediated transformation.
Claim 17 limits the site specific nuclease to Cas9 and so on.
Regarding claims 1, 3, 17
Shultz et al teach a method of altering/editing a genome of a plant comprising:
delivering construct comprising DNA encoding RNA-guide site-specific nuclease (Cas9) into plant (nicotiana benthamiana and maize) seeds comprising embryo (Examples 4-6, [00103]-[00122]).
Dry mature seeds were suspended (thus are removed from the plant) and wetted in MS culture medium before transformation ([00107]), reading on explant. The cultured and wetted seed reads on dry mature plant embryo explant.
Shultz et al teach and demonstrated that “each test or control treatment experiment is carried out on 10 dry excised embryos” (Example 5 in [00113]). Such dry excised embryos read on "dry-excised mature plant embryo explant" defined by instant specification.
The construct comprises plant expressible promoter (PcUbi4-2, Example 4, table 5, [00105]).
After the transformation, the seeds comprising genomic modification is growing and producing progeny, thus, the explants were regenerated to a plant. The plant and progeny were tested for gene modification using phenotype and/or molecular assay. The seed, plant and progeny comprise editing and gene integration (each of the Examples 4-6, [00111], [00115], [00121]).
Thus, Shultz et al teach and demonstrated all of the limitations of claims 1, 3 and 17,
except
1. do not teach the step of excising an embryo from a dry mature plant seed, thereby producing a dry-excised mature plant embryo explant (the new limitation in the amendment), and
2. do not explicitly teach the transformation is via bacterially mediated transformation to delivery the recombinant construct (Examples 4-6 are not using bacterially mediated transformation, thus, a 102 rejection is not made).
Regarding 1., as analyzed Shultz et al teach and demonstrated using "dry-excised mature plant embryo explant" for the transformation/delivery (Example 5 in [00113]). Thus, Shultz et al provide a strong motivation to perform the step.
Patnaik et al teach and demonstrate excising embryos from sterilized dry mature seeds (p308, left col, 1st para in “Materials and Methods”), and teach using the embryo (reading on "dry-excised mature plant embryo explant") for Agrobacterium-mediated transformation (p308, left col, last para, right col, 1st para, in “Materials and Methods”). Patnaik et al demonstrated successful transformation (Results in p310-312).
Regarding 2., Shultz et al nevertheless suggest that Agrobacterium mediated transformation is a routine technique and is an embodiment of the invention including using Agrobacterium mediated transformation to deliver CRISPR-Cas nucleases ([0020], [0023], [0025], [0035]). Shultz et al also cite references teaching Agrobacterium mediated transformation ([0020]).
Li et al teach a detailed method of transforming soybean by Agrobacterium mediated transformation (Title, Abstract, whole article).
Specifically, Li et al teach imbibing mature and dry soybean seeds (reading on dry mature plant embryo explant), and culturing the imbibed seeds with Agrobacteria (p4, left col, last para, right col, 1st to 2nd paras).
Li et al demonstrated that the transformation was successful, and transgenic plants were regenerated from the infected seeds (p4, fig 2, J and K, Results in p6-11).
Li et al teach that Agrobacteria and particle bombardment are functional equivalent in plant transformation, and that about 85% of the transgenic plants are obtained using the Agrobacterium-mediated transformation method (p2, left col, 2nd and 3rd para).
Hence, the only difference between Shultz et al and Li et al is a nuclease vs another DNA molecule of interest.
In addition to Patnaik et al and Li et al, Char et al teach and demonstrated transforming/delivering Cas9 gene into plant (maize) cells to produce transgenic and mutant maize seeds comprising Cas9 gene by Agrobacterium-mediated transformation (p257, Abstract; Result, p259, left col, 1st para, figure 2). Hence, Char et al support Patnaik et al and Li et al, and provide added expectation of success.
Regarding dependent claims
Shultz et al teach that the transformation used a construct comprising T-DNA (Same Example 4, [00104], claim 8), the limitation of claim 4.
Shultz et al teach that the gene of interest/insertion/transgene and the nuclease are in the same construct (example 4, [00104], table 5), thus, the edit or site-specific integration is near the site of nuclease, the limitation of claim 12.
As analyzed above, after the transformation, the plant and progeny were tested for gene modification using phenotype and/or molecular assay. The seed, plant and progeny comprise editing and gene integration (Example 4, [00103], [00111]). Thus, Shultz et al teach the limitation of claim 13.
Shultz et al suggest Cpf1, like Case9, is a Cas endonuclease, and can be transformed to the dry mature plant embryo explant ([0012], [0015], [0017], claim 18), the limitation of claim 18.
Shultz et al teach that guide-RNA and terminators and the nuclease are in the same construct (example 4, [00104], table 5). Claim 1 only requires nuclease. The guide RNA or terminator is a second recombinant DNA molecule. Thus, Shultz et al teach the limitation of claim 23.
Regarding claims 25-26, 48-50, according to specification ([0083]), a “donor molecule”, “donor template”, or “donor template molecule” (collectively a “donor template”), which may be a recombinant polynucleotide, DNA or RNA donor template or sequence, is defined as a nucleic acid molecule having a homologous nucleic acid template or sequence (e.g., homology sequence) and/or an insertion sequence for site-directed, targeted insertion or recombination into the genome of a plant cell via repair of a nick or double-stranded DNA break in the genome of a plant cell. Shultz et al teach that the construct comprises multiple insertions including gene of interest (encoding aminoglycoside adenylyltransferase sequences for selection, reading on transgene and marker gene) and promoter (Example 4, [00104]-[0105], table 5). The construct comprises CRISPR-Cas ([00105]) and transgene, which produces site-specific genome modification by homologous recombination ([0028]). Accordingly, Shultz et al teach second DNA molecule (the first is nuclease) being donor molecule along with the nuclease encoding molecule, the molecule being an insertion, a transgene/marker gene, teaching the limitations of claims 25-26, and 48-50.
Regarding claims 39-40, 47, the specification uses “construct” and “vector” interchangeably ([0101]). Schultz et al teach delivering a second DNA molecule thus teach claim 23, but do not teach delivering a second DNA construct comprising guide RNA in Example 4 above. Schultz et al, however, teach that guide RNA expressed from one vector/construct, and the RNA guided nuclease from a second vector. Schultz et al also teach using different vectors/constructs for expressing different guide RNAs ([0021]). Schultz et al actually made different vectors to express nuclease and guide RNA (sgRNA) (Example 8, [00135]). Thus, Shultz et al at least suggest the limitation of claims 39-40, 47.
Nicotiana benthamiana above in Example 4 is a dicot, thus, the Shultz et al teach the limitation of claim 60.
Shultz et al applying the method to soybean ([0039]) and the method of editing gene of soybean (Example 1, [0073]), the limitation of claim 61.
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 Shultz et al, such that the method includes a step of excising an embryo from a dry mature plant seed, as motivated by Shultz et al, and taught and demonstrated by Patnaik et al. It also would have been obvious for one ordinary skill in the art to modify the invention of Shultz et al, such that the method includes using Agrobacterium mediated transformation to deliver the site-specific nuclease Cas9, as strongly suggested by Shultz et al, and explicitly taught and demonstrated by Li et al, and Char et al. One ordinary skill in the art would have been motivated to do so, because Agrobacterium mediated transformation is a functional alternative of other transformation methods and demonstrated in detail by Li et al and Char. The expectation of success would have been high, because both Shultz et al and Li et al used the same imbibed dry mature plant embryo explant, the only difference between Shultz et al and Li et al is a nuclease vs another DNA molecule of interest, and the Agrobacterium mediated transformation is a routine method producing 85% of transgenic plants. In addition, Char et al support Patnaik et al and Li et al, and provide added expectation of success.
Furthermore, as analyzed above, the specification does not provide any example of “delivering to a dry mature plant embryo explant via bacterially mediated transformation a recombinant DNA construct”. Thus, the applicant merely claims what the prior art taught or/and suggested.
Therefore, the invention would have been obvious to one ordinary skill in the art.
Claim 62 is rejected under 35 U.S.C. 103 as being unpatentable over Shultz et al in view of Patnaik et al, Li et al and Char et al, as applied to claim 1 above, and further in view of Liang et al (Selection of highly efficient sgRNAs for CRISPR/Cas9-based plant genome editing. Nature Scientific Reports, 1-8, 2016).
Claim 1 has been analyzed above.
Claim 62 limits claim 1, wherein the dry-excised mature plant embryo explant comprises, prior to the delivering step, a guide RNA (gRNA).
As analyzed above, Schultz et al teach that the guide RNA and the DNA encoding the site-specific nuclease are in different vectors. But Schultz et al do not teach delivering DNA encoding site-specific nuclease to plant cells comprising guide RNA (prior to delivery).
Liang et al teach that some guide RNAs comprised in plant cell could lead to off-target editing by CRISPR-Cas (not at intended site, thus not desirable), and teach locating (intended and unintended) guide RNAs (sgRNA) in plant cells for next testing (p4, 2nd para). Liang et al continue to teach the next testing is to deliver Cas-sgRNA (Cas is a site-specific nuclease) to the plant cells comprising the (intended and unintended) guide RNAs, for selecting guide-RNAs leading to predicted/intended target sites (p4, last para; p5, 1st para).
Liang et al further teach the advantage of such tests is reducing off-targeting and increasing editing accuracy (p4, 2nd para; p6, 1st para).
Thus, Liang et al provide a motivation to perform such tests and selections.
It would have been obvious for one ordinary skill in the art to modify the invention of Shultz et al, such that the method further comprises delivering Cas-sgRNA (Cas is a site-specific nuclease) to the plant cells comprising guide RNA, as suggested by Shultz et al and motivated by Liang et al. One ordinary skill in the art would have been motivated to do so, because of the advantage of testing guide-RNAs and selecting that are specific to the intended editing from plant cells as taught and demonstrated by Liang et al. The expectation would have been high, because the method steps of Schultz et al and Liang et al are essentially the same and routine, only difference is the target plant cells comprise guide RNAs. Off-target testing is also routine as taught by Liang et al.
Therefore, the dependent claim would have been obvious to one ordinary skill in the art.
Claim 65 is rejected under 35 U.S.C. 103 as being unpatentable over Shultz et al in view of Patnaik et al, Li et al, and Char et al, as applied to claim 1 above, and viewed with evidence from Tri-States (Dry-Grain Management Means Longer Storage Times for Wheat, 8/20/2018).
Claim 1 has been analyzed above.
Claim 65 limits claim 1, wherein the dry-excised mature plant embryo explant has a moisture content within a range from about 3% to about 25% or wherein the dry mature embryo explant is excised from a plant seed having a moisture content within a range from about 3% to about 25%.
Shultz et al in view of Patnaik et al, Li et al, and Char et al, are silent on such a specific limitation.
Patnaik et al further teach excising an explant from dry mature (wheat) plant seed, (307, right col, last para, p308, left col, 1st para), and used the explant for transformation (p308, right col, 1st para) and demonstrated success by PCR and protein assay (p309, whole page).
Tri-States provides evidence that the ideal wheat moisture content at harvest is between 18% and 20%. This is above the ideal wheat moisture content for stored wheat. You’ll want to bring the moisture content down to 13.5% if you plan to sell soon, since that is the ideal wheat moisture content for the best-selling price. If you plan to store the wheat for a year, or more, you’ll want to bring it down even more to 12.5%.
Thus, the mature wheat seed of Patnaik et al reads on “the mature embryo explant is excised from a plant seed having a moisture content within a range from about 3% to about 25%”, the limitation of claim 65.
Therefore, the dependent claim would have been obvious to one ordinary skill in the art as claim 1 would have been.
Remarks
For compact prosecution, the following reference is relevant to instant applicant, thus, is filed but not cited by the examiner:
Paz et al (Improved cotyledonary node method using an alternative explant derived from mature seed for efficient Agrobacterium-mediated soybean transformation. Plant Cell Rep. 25: 206–213, 2006).
Response to Arguments
Double Patenting
The terminal disclaimer of 8/26/2026 has been approved, overcoming the rejection.
35 USC § 103
Claims have been significantly amended. The rejections are modified accordingly. Patnaik et al, cited in the rejections, was cited previously for claim 65. Char et al is a new reference. Thus, the arguments regarding the previous rejections are mostly not applicable. Nevertheless, the brief responses are provided below.
The applicant argues that No claim is obvious because neither Shultz nor Li teaches or suggests excising an embryo from a dry mature seed.
The argument is fully considered but is not deemed persuasive particularly to the instant rejections. Shultz et al do not teach the excising step, but teach and demonstrated using a "dry-excised mature plant embryo explant" for the transformation/delivery (Example 5 in [00113]). Thus, Shultz et al provide a strong motivation to perform the step.
Nevertheless, Patnaik et al explicitly teach such a step, and demonstrated using the resultant embryo explant to successfully transform plant cells via Agrobacterium as a medium, as analyzed above.
Char et al is cited as an additional support and provide additional expectation of success.
Therefore, such step and the claims reciting the step are deemed obvious.
In addition, as analyzed above, the specification does not provide any example of “delivering to a dry mature plant embryo explant via bacterially mediated transformation a recombinant DNA construct”. Thus, the applicant merely claims what the prior art taught or/and suggested.
Regarding dependent claims 62 and 65, their patentability depends on that of the parent claim. In addition, 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. Any suggestion or motivation can be used in a 103 rejection.
Regarding claim 62, Liang et al teach that some guide RNAs comprised in plant cell could lead to off-target editing by CRISPR-Cas (not at intended site, thus not desirable), and teach locating (intended and unintended) guide RNAs (sgRNA) in plant cells for next testing (p4, 2nd para). Liang et al continue to teach the next testing is to deliver Cas-sgRNA (Cas is a site-specific nuclease) to the plant cells comprising the (intended and unintended) guide RNAs, for selecting guide-RNAs leading to predicted/intended target sites, and further teach the advantage of such tests is reducing off-targeting and increasing editing accuracy. Thus, Liang et al provide a motivation to perform such tests and selections.
Regarding 65, Tri-States provides evidence that the ideal wheat moisture content at harvest is between 18% and 20%, and that the mature wheat seed of Patnaik et al reads on the claim limitation.
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.
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/Wayne Zhong/
Primary Examiner, Art Unit 1662
