DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application is being examined under the pre-AIA first to invent provisions.
Election/Restrictions
Applicant’s election without traverse of Group I, claim 13, in the reply filed on May 13, 2026 is acknowledged. Claim 12 links inventions I-IX. Claims 14-21 are withdrawn.
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of pre-AIA 35 U.S.C. 103(a) which forms the basis for all obviousness rejections set forth in this Office action:
(a) A patent may not be obtained though the invention is not identically disclosed or described as set forth in section 102, if the differences between the subject matter sought to be patented and the prior art are such that the subject matter as a whole would have been obvious at the time the invention was made to a person having ordinary skill in the art to which said subject matter pertains. Patentability shall not be negatived by the manner in which the invention was made.
This application currently names joint inventors. In considering patentability of the claims under pre-AIA 35 U.S.C. 103(a), the examiner presumes that the subject matter of the various claims was commonly owned at the time any inventions covered therein were made absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and invention dates of each claim that was not commonly owned at the time a later invention was made in order for the examiner to consider the applicability of pre-AIA 35 U.S.C. 103(c) and potential pre-AIA 35 U.S.C. 102(e), (f) or (g) prior art under pre-AIA 35 U.S.C. 103(a).
Claim s 12 and 13 is/are rejected under pre-AIA 35 U.S.C. 103(a) as being unpatentable over Yoshikuni et al. U.S. Patent Application Publication No. 2019/0048354, published Feb. 14, 2019.
Claim 12 is drawn to a method for inserting at least two different polynucleotides of interest into the genome of a host cell, the method comprising the steps of:
a) providing a host cell comprising in its genome:
i. at least two polynucleotides encoding at least two different selectable markers, each comprising a different target sequence flanked by a functional PAM sequence for a Class-II Cas9 protein;
ii. at least two polynucleotides encoding at least two gRNAs that are at least 80% complementary to and capable of hybridizing to the at least two different target sequences;
iii. a polynucleotide encoding a nuclease-null variant of a Class-II Cas9 protein capable of interacting with the at least two gRNAs and binding to the at least two different target sequences, whereby expression of the two different selectable markers is repressed;
b) transforming said host cell with at least two different polynucleotides of interest, said
polynucleotides being capable of inactivating the at least two polynucleotides encoding the at
least two gRNAs; and
c) selecting for the traits conferred by the at least two different selectable markers; and
d) identifying a transformed host cell, wherein the at least two polynucleotides encoding the at least two gRNAs have been inactivated by the at least two different polynucleotides of interest.
Claim 13 is drawn to the method according to claim 12, wherein the at least two polynucleotides of interest encode an enzyme independently selected from hydrolase, isomerase, ligase, lyase, oxidoreductase, or transferase.
Yoshikuni et al. teach a method for inserting a polynucleotide of interest into the genome of a host cell, the method comprising the steps of providing a host cell comprising in its genome a polynucleotide encoding a first selectable marker flanked by recombinase target sequences, transforming the host cell with a polynucleotide of interest comprising a gene of interest encoding PDC core pathway enzymes and a second selectable marker coding sequence flanked by recombinase target sequences, and also encoding an inducible recombinase whose expression is repressed, inducing the expression of the recombinase, selecting for the trait conferred by the second selectable marker, and identifying a transformed host cell, wherein the polynucleotide encoding the first selectable marker has been inactivated by the polynucleotide of interest (Example 1 paragraphs [0143] to [0154]; claim 1). The polynucleotide of interest can also comprise coding sequences for other types of enzymes, including enzymes that catalyze one or more steps in the synthesis of a polysaccharide (claims 39-40), which inherently includes at least transferase and ligase enzymes (see, e.g. Woodward et al. Bacterial polysaccharide synthesis and export. Current Opinion in Structural Biology. Volume 40, October 2016, Pages 81-88).
Yoshikuni et al. also teach that it is known in the art that CRISPR-Cas9 refers to a two component ribonucleoprotein complex with guide RNA and a Cas9 endonuclease, and that in type II systems, the guide RNA interacts with Cas9 and directs the nuclease activity of Cas9 to target DNA sequences complementary to those present in the guide RNA, where the guide RNA base pairs with complementary sequence in target DNA (paragraph [0076]).
Yoshikuni et al. additionally teach that the guide target sequence is complementary to the guide RNA target sequence, and that any desired target DNA sequence of interest may be targeted by a guide RNA target sequence, provided that it meets the requirement for the presence of a protospacer-adjacent motif (PAM) adjacent to the sequence complementary to the sgRNA target sequence (paragraph [0084]).
Yoshikuni et al. further teach that the CRISPR/Cas9 system has been engineered to promote repression or activation of gene transcription, with the CRISPRi system having been developed to repress gene transcription through a catalytically dead Cas9 (dCas9) protein/guide RNA complex, where the catalytically dead Cas9 (dCas9) protein lacks endonucleolytic activity but retains the ability to bind DNA in a site-specific manner targeted by the complexed guide RNA (paragraph 0078]).
Yoshikuni et al. also teach transformation methods that employ a third selectable marker coding sequence (claims 148-149 and 156-157).
Yoshikuni et al. do not teach the use of a polynucleotide encoding a selectable marker comprising a target sequence flanked by a functional PAM sequence for a Class-Il Cas9 protein in conjunction with at least one polynucleotide encoding a gRNA that is at least 80% complementary to and capable of hybridizing to the target sequence a polynucleotide encoding a nuclease-null variant of a Class-II Cas9 protein capable of interaction with the gRNA and binding to the target sequence, whereby expression of the selectable marker is repressed and a polynucleotide of interest that is capable of inactivating the at least one polynucleotide encoding the gRNA.
Given the teachings of Yoshikuni et al. that a host cell transformed with a polynucleotide of interest can be identified by selecting for a trait conferred by a second selectable marker when a polynucleotide of interest comprising a gene of interest encoding one or more enzymes, a second selectable marker coding sequence flanked by recombinase target sequences, and an inducible recombinase coding sequence, is introduced into a host cell comprising a first selectable marker gene flanked by recombinase target sequences whereby the first selectable marker gene is inactivated by recombination, given the teachings of Yoshikuni et al. that a CRISPR/Cas9 system has been engineered to promote the repression of gene transcription (CRISPRi) through a catalytically dead Cas9 (dCas9) protein/guide RNA complex that retains the ability to bind DNA in a site-specific manner targeted by the complexed guide RNA, and given the further teachings of Yoshikuni et al. that transformation methods may employ multiple selectable marker coding sequences, it would have been prima facie obvious to a person having ordinary skill in the art before the effective filing date of the claimed invention that a host cell transformed with one or more polynucleotides of interest can also be identified by selecting for one or more traits conferred by one or more selectable marker genes comprising one or more target sequences flanked by a functional PAM sequence for a Class-Il Cas9 protein in a host cell that expresses a nuclease-null variant of a Class-II Cas9 protein and one or more gRNAs capable of hybridizing to the target sequences to repress the expression of the selectable marker genes when the one or more polynucleotides of interest introduced into a host cell inactivate the gRNA coding sequences by recombination.
One skilled in the art would have recognized that the inactivation of the gRNA coding sequences by the polynucleotides of interest would derepress the selectable marker genes and allow for the selection of the host cell transformed with the polynucleotides of interest, since the prior art recognizes that the dCas9 repressor protein only binds a DNA target sequence when the dCas9 is complexed with a gRNA specific for the target sequence. The use of one or more polynucleotides of interest that derepress selectable marker genes present in a host cell by recombinationally inactivating a repressor component in place of a polynucleotide of interest that recombinationally replaces a first selectable marker gene present in a host cell with a second selectable marker gene would have been a simple substitution of equivalent elements to obtain predictable results, i.e. the selection of the host cell transformed with one or more polynucleotides of interest.
Thus, the claimed invention would have been prima facie obvious as a whole to a person having ordinary skill in the art before the effective filing date of the claimed invention.
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on nonstatutory double patenting provided the reference application or patent either is shown to be commonly owned with the examined application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. See MPEP § 717.02 for applications subject to examination under the first inventor to file provisions of the AIA as explained in MPEP § 2159. See MPEP § 2146 et seq. for applications not subject to examination under the first inventor to file provisions of the AIA . A terminal disclaimer must be signed in compliance with 37 CFR 1.321(b).
The USPTO Internet website contains terminal disclaimer forms which may be used. Please visit www.uspto.gov/patent/patents-forms. The filing date of the application in which the form is filed determines what form (e.g., PTO/SB/25, PTO/SB/26, PTO/AIA /25, or PTO/AIA /26) should be used. A web-based eTerminal Disclaimer may be filled out completely online using web-screens. An eTerminal Disclaimer that meets all requirements is auto-processed and approved immediately upon submission. For more information about eTerminal Disclaimers, refer to www.uspto.gov/patents/process/file/efs/guidance/eTD-info-I.jsp.
Claims 12 and 13 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claim 16 and 17 of copending Application No. 19/430,247 (reference application).
Although the claims at issue are not identical, they are not patentably distinct from each other. The method of claim 12 of the instant application only differs from the method of claim 16 of the reference application in that the method of claim 12 of the instant application utilizes a polynucleotide encoding a nuclease-null variant of a Class-II Cas9 protein, whereas the method of claim 16 of the reference application utilizes a polynucleotide encoding a nuclease-null variant of an RNA-guided endonuclease, making the method of claim 12 of the instant application a species of the method of claim 16 of the reference application. Further, the method of claim 13 of the instant application differs from the method of claim 17 of the reference application in that the method of claim 13 of the instant application utilizes a specific set of enzymes, whereas the method of claim 17 of the reference application only utilizes a generic enzyme, making the method of claim 13 of the instant application a set species of the method of claim 17 of the reference application.
This is a provisional nonstatutory double patenting rejection because the patentably indistinct claims have not in fact been patented.
Remarks
Any inquiry concerning this communication or earlier communications from the examiner should be directed to CYNTHIA E COLLINS whose telephone number is (571)272-0794. The examiner can normally be reached M-F 8:30 am - 5:00 pm.
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/CYNTHIA E COLLINS/Primary Examiner, Art Unit 1662
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