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 .
Application Status and Withdrawn Rejections
Applicant’s amendment filed November 26, 2025, amending claims 10 and 24 is acknowledged. Claims 1-4 and 6-26 are pending. Claim 19 remains withdrawn from further consideration pursuant to 37 CFR l.142(b) as being drawn to a nonelected group, there being no allowable generic or linking claim.
Claims 1-4, 6-18 and 20-26 are under examination.
The claim amendments overcome the §112(b) rejection. Any other rejection or objection not reiterated herein has been overcome by amendment. Applicant’s amendments and arguments have been thoroughly reviewed, but are not persuasive to place the claims in condition for allowance for the reasons that follow.
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.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Claims 1-4, 6, 8-10, 12-13, 15-18 and 20-26 are rejected under 35 U.S.C. 103 as being unpatentable over DiCarlo (DiCarlo et al., Nucleic Acids Research (2013), 41: 4336-4343; of record) in view of Zhang II (US 20140357530 A1, published December 4, 2014; of record), Wagner (Wagner et al., Nature Methods (2014), 11: 915-918; of record) and Benton (Benton et al., Molecular and Cellular Biology (1990), 10: 353-360; of record). This is a maintained rejection.
Regarding claims 1, 20-21 and 25-26, DiCarlo teaches using Cas9 and engineered guide RNAs (gRNAs) for genome editing in the yeast Saccharomyces cerevisiae (Abstract). DiCarlo teaches transforming (i.e., inserting into) S. cerevisiae cells a plasmid encoding the gRNA (i.e., a polynucleotide encoding the guide RNA) (page 4337, ¶4). DiCarlo teaches the gRNA polynucleotide is operably linked to the SNR52 promoter (page 4338, ¶2; Fig 1). DiCarlo teaches the SNR52 promoter is a snoRNA promoter, which is an RNA polymerase III promoter (page 4339, ¶4).
DiCarlo does not teach driving gRNA expression from a viral single-subunit DNA-dependent RNA polymerase promoter with transcription performed by the single-subunit DNA-dependent RNA polymerase.
Zhang II teaches using a T7 promoter (i.e., a viral single-subunit DNA-dependent RNA polymerase) to drive the expression of guide RNAs ([0019] and [0468]). Zhang II also teaches delivering to eukaryotic cells expression vectors (i.e., polynucleotides) for the T7 polymerase and the guide RNA fused (i.e., operably linked) to the T7 promoter ([0468] - [0472]). Zhang II expressly teaches "[r]ather than use pol3 promoters, in particular RNA polymerase Ill (e.g. U6 or HI promoters), to express guide RNAs in eukaryotic cells, Applicants express the T7 polymerase in eukaryotic cells to drive expression of guide RNAs using the T7 promoter." ([0468]).
Wagner demonstrates the functionality of expressing T7 RNA Polymerase (RNAP) in the single celled eukaryote, Plasmodium falciparum, and driving Cas9 gRNA expression from the T7 promoter (Figure 2). Wagner teaches choosing the T7 promoter for gRNA expression because the T7 RNAP uses well-characterized promoters to make transcripts of defined size and in high yield (page 915, ¶2).
Benton teaches expressing T7 RNAP in S. cerevisiae cells (Abstract; page 355, ¶4). Benton teaches T7 RNAP transcribed RNAs from the T7 DNA f10 promoter (i.e., a T7 promoter) (page 356, ¶5). Benton teaches upon induction of T7 RNAP, “substantial amounts of target RNA accumulated” (page 356, ¶6).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have substituted the SNR52 promoter for Cas9 guide RNA expression in the S. cerevisiae gene editing system of DiCarlo for the T7 RNAP-promoter based expression system for Cas9 gRNAs as taught in Zhang II and Wagner. It would have amounted to a simple substitution of one known promoter for the expression of a Cas9 gRNAs for another by known means to yield predictable results. Zhang suggests using the T7 promoter to drive gRNA expression in a generic cell, while Wagner demonstrates the success of using the T7 promoter to drive gRNA expression in a single-celled eukaryote. The skilled in the art would have had a reasonable and predictable expectation of success that T7 promoter could also drive expression of a gRNA in S. cerevisiae because Benton demonstrates that T7 RNAP can be expressed in S. cerevisiae and drives expression of target RNAs from its T7 promoter. The skilled artisan would have been motivated to replace the SNR52 promoters of DiCarlo with the T7 expression system because both Wagner and Benton teaches target RNAs are produced in high and substantial amounts from the T7 promoter.
Regarding claim 2 and 22, Benton teaches expressing the RNAP from both a plasmid (i.e., a vector) and from the yeast chromosome (i.e., from the genome) (page 355, ¶4). Benton teaches expressing T7 RNAP from a multi-copy plasmid comprising the LEU2 marker (i.e., a selectable marker) (page 354, ¶2).
Regarding claim 3, DiCarlo teaches the gRNA is expressed from a plasmid (i.e., a vector) (page 4338, ¶5) or as a transient gRNA cassette (i.e., a linear nucleic acid construct) (page 4338, ¶4).
Regarding claim 4, DiCarlo teaches the gRNAs are from a CRISPR/Cas9 (i.e., CRIPSR/Cas) nuclease system (Fig 1).
Regarding claim 6, Benton teaches the T7 RNAP is expressed from the GAL1-GAL10 galactose-inducible promoter (page 654, ¶2; Fig 1).
Regarding claims 8 and 23, Benton teaches the T7 RNAP comprises a nuclear localization signal from SV40 at the N-terminus (Table 1).
Regarding claim 9-10, DiCarlo teaches expressing 4 different gRNAs in S. cerevisiae (Fig 1B, 2C, 3B). Benton teaches driving lac, cat and lacZ RNAs (i.e., multiple RNAs) from the T7 promoter (Figs 3-4). Zhang II teaches expressing two DR-crRNA arrays targeting two separate loci (i.e., multiple, distinct guide-RNAs) from the same promoter in a cell (Figure 4F). Zhang II teaches libraries of guide RNAs against a plurality of target sequences for the purpose of creating a libraries of knock cells in used in functional genomics ([0010-0011]).
It also would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have expressed multiple guide RNAs as taught in DiCarlo and Zhang II under the control of one or more T7 promoters because it would have amounted to a simple combination of known guide RNAs and promoters by known means to yield predictable results. As indicated by both Zhang II, it is well known in the art that multiple guide RNAs can be simultaneously expressed in a cell for the simultaneous editing of multiple targets. Thus, one skilled in the art would have a reasonable and predictable expectation of success of expressing multiple distinct guide RNAs under the control of one or more T7 promoters. One would have been motivated to do so in order to edit two targets either for complete gene knockout or to discover synthetic lethal gene interactions.
Regarding claim 12, Wagner teaches using a T7 terminator for transcriptional termination of the T7-driven gRNA (page 915, ¶2; Online Methods, page 2, ¶1). Wagner teaches using the T7 terminator allows the cell to make transcripts of a defined size (page 915, ¶2). Benton teaches using the Tf transcription terminator for T7 DNA (i.e., a viral single-subunit DNA-dependent RNA polymerase terminator) in combination with the T7 promoter in S. cerevisiae (page 358, ¶2). Benton teaches that T7 RNA polymerase recognizes its specific termination signal in yeast nuclear DNA (page 358, ¶2).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have also included the T7 terminator taught in Wagner and Benton in the T7 promoter driven gRNA expression cassette. It would have amounted to the simple combination of known regulatory elements by known means to yield predictable results. The skilled artisan would have predicted that the T7 terminator could be used in the T7p-gRNA construct specifically in yeast because Benton teaches that T7 RNAP recognizes its terminator in yeast. The skilled artisan would have been motivated to do so because both Wagner and Benton teach using the T7 terminator allows RNAs of specific length to be produced.
Claim 13 recites "wherein the polynucleotide and viral single-subunit DNA-dependent RNA polymerase promoter are present on a plasmid, and wherein the plasmid is assembled within the cell by integration of a single-stranded or double-stranded oligonucleotide comprising the target sequence of the guide-polynucleotide into the plasmid". The broadest reasonable interpretation of this clause is as a product-by-process limitation. This product-by-process will be examined according to the structure of the plasmid that is produced by integrating a single-stranded or double-stranded oligonucleotide comprising the target sequence of the guide-polynucleotide into the plasmid. The specification (page 18) and incorporated reference EP 16181781.2 (priority document to US Patent 11,149,268 B2) indicate that the plasmid contains a 20-nt target sequence of a guide RNA operably linked upstream of the structural component of the guide RNA and downstream of a promoter. DiCarlo teaches the gRNA is downstream of the SNR52 promoter (Fig 1B). Wagner teaches the coding sequence for the gRNA is downstream of the T7 promoter (Fig 2A).
Regarding claim 15, DiCarlo teaches S. cerevisiae cells also expressing Cas9 which is functional for genome editing (Figs 1B, 2C). DiCarlo teaches Cas9 is a type II bacterial CRISPR system of Streptococcus pyogenes (page 4336, ¶1). Because DiCarlo teaches that Cas9 naturally occurs in bacteria (page 4336, ¶1), Cas9 is heterologous to DiCarlo’s S. cerevisiae cells.
Regarding claims 16-18, the teachings of DiCarlo, Zhang II, Wagner and Benton, and the obviousness of using the T7 promoter and T7 RNAP to express the Cas9 guide RNA of DiCarlo in S. cerevisiae cells are recited above as for claim 1. The product of the method rendered obvious above would produce a S. cerevisiae cell comprising the T7 RNAP and the T7 promoter operably linked to the guide RNA encoding polynucleotide, which reads on the composition of claim 16 and the cells of claims 17 and 18.
Regarding claim 24, the Specification indicates that a compound of interest can be a polypeptide (page 4). DiCarlo teaches the cells comprising the T7 promoter-driven gRNA and the T7 RNAP can also express Cas9 (i.e., a polypeptide, a compound of interest), as evidenced by the mutation rate of the targeted DNA (Fig 2C-D).
Claim 7 is rejected under 35 U.S.C. 103 as being unpatentable over DiCarlo (DiCarlo et al., Nucleic Acids Research (2013), 41: 4336-4343; of record), Zhang II (US 20140357530 A1, published December 4, 2014; of record), Wagner (Wagner et al., Nature Methods (2014), 11: 915-918; of record) and Benton (Benton et al., Molecular and Cellular Biology (1990), 10: 353-360; of record) as applied to claims 1-4, 6, 8-10, 12-13, 15-18 and 20-26 above, and further in view of Enayati (Enayati et al., FEMS Microbiology Letters (2016), 363(5); 1-7, published February 5, 2016; of record). This is a maintained rejection.
The teachings of DiCarlo, Zhang II, Wagner and Benton are recited above and applied as for claims 1-4, 6, 8-10, 12-13, 15-18 and 20-26. Zhang II also teaches codon-optimization of bacterial genes for expression in eukaryotic cells ([0025, [0059]). Zhang teaches that codon optimization methods of host species is known ([0025]).
DiCarlo, Zhang II, Wagner and Benton do not teach codon-optimization of the T7 RNAP.
Enayati teaches "T7-based expression systems have been broadly used for in vivo or in vitro generation of ssRNA and dsRNA." (Page 4, ¶4). Enayati teaches a system for expressing non-coding RNAs using a T7 promoter and T7 RNA polymerase in the fungus Aspergillus fumigatus (Abstract; one sentence summary). Enayati teaches the gene sequence encoding T7 RNAP is codon-optimized for expression in the experimental fungal cell model (page 2, ¶5).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have encoded the T7 RNAP using codons recognized by S. cerevisiae. It would have amounted to applying a known method of heterologous gene expression by known means to yield predictable results. The skilled artisan would have predicted that the coding sequence for RNAP could be optimized for expression in S. cerevisiae because Enayati demonstrates codon optimization of RNAP in a different fungal species and Zhang II teaches that methods of codon optimization are known in the art. The skilled artisan would have been motivated to do so to optimize expression of RNAP in S. cerevisiae.
Claim 11 is rejected under 35 U.S.C. 103 as being unpatentable over DiCarlo (DiCarlo et al., Nucleic Acids Research (2013), 41: 4336-4343; of record), Zhang II (US 20140357530 A1, published December 4, 2014; of record), Wagner (Wagner et al., Nature Methods (2014), 11: 915-918; of record) and Benton (Benton et al., Molecular and Cellular Biology (1990), 10: 353-360; of record) as applied to claims 1-4, 6, 8-10, 12-13, 15-18 and 20-26 above, and further in view of Jones (Jones et al., Sci Rep (2015) 5, 11301; Published June 11, 2015, of record). This is a maintained rejection.
The Specification defines variant promoters that have a difference sequence as compared to the wild-type promoter found in nature while still retaining promoter activity (page 14).
The teachings of DiCarlo, Zhang II, Wagner and Benton are recited above and applied as for claims 1-4, 6, 8-10, 12-13, 15-18 and 20-26.
DiCarlo, Zhang II, Wagner and Benton do not teach or suggest the T7 RNA Polymerase promoter is variant viral DNA-dependent RNA polymerase promoter.
Jones teaches developing and characterizing a library of T7 promoters (i.e., a library of single subunit viral DNA-dependent RNA polymerase promoters) (page 3, ¶2). Jones also teaches the mutant (i.e., variant) T7 promoters from the library varied in the ability to express a fluorescent reporter protein (Figure 2). Jones teaches that some T7 mutant promoters are stronger than wild-type, while other mutants were weaker than wild type (Figure 2). Jones teaches that using promoters of varying strength can result in more desired outcomes and that using the strongest or weakest promoters does not always produce the best results (Figure 3). Jones teaches several T7 promoter mutants that either increase or decrease expression of a fluorescent reporter molecule relative to the wild type promoter (Figure 2).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have expressed multiple guide RNAs as taught in DiCarlo and Zhang II under the control of two or more variant T7 promoters discovered in the library of Jones. It would have amounted to a simple substitution of known T7 promoters for others by known means to yield predictable results. Jones teaches that some of the T7 promoter discovered in the library increase expression or decrease expression of the transgenes, relative to wild type T7 promoter. One skilled in the art would predict that variant T7 promoters would also drive expression in S. cerevisiae cells since Benton teaches wild type T7 promoter is functional in S. cerevisiae. One would be motivated to make the substitution in order to increase or decrease the expression of the different guide RNAs depending on the expression level of the Cas9 nuclease and the level of gene modification desired.
Claim 13 is rejected under 35 U.S.C. 103 as being unpatentable over DiCarlo (DiCarlo et al., Nucleic Acids Research (2013), 41: 4336-4343; of record), Zhang II (US 20140357530 A1, published December 4, 2014; of record), Wagner (Wagner et al., Nature Methods (2014), 11: 915-918; of record) and Benton (Benton et al., Molecular and Cellular Biology (1990), 10: 353-360; of record) as applied to claims 1-4, 6, 8-10, 12-13, 15-18 and 20-26 above, and further in view of van Leeuwen (van Leeuwen et al., Cold Spring Harb Protoc (2015) 2015(9), 853-861; published September 1, 2015; of record). This is a maintained rejection.
As recited above in paragraph 23, the broadest reasonable interpretation of “wherein the polynucleotide and single-subunit DNA-dependent RNA polymerase promoter are present on a plasmid, and wherein the plasmid is assembled within the cell by integration of a single-stranded or double-stranded oligonucleotide comprising the target sequence of the guide-polynucleotide into the plasmid" is as a product-by-process limitation. However, in the event that the claim is amended to include the clause as steps in the method, the following rejection would apply.
The teachings of DiCarlo, Zhang II, Wagner and Benton are recited above and applied as for claims 1-4, 6, 8-10, 12-13, 15-18 and 20-26.
DiCarlo, Zhang II, Wagner and Benton do not teach plasmid assembly in yeast cells.
Van Leeuwen teaches a method of assembling plasmids in a yeast cell using homologous recombination of double stranded plasmid parts (Figure 1). Van Leeuwen teaches the parts have 30 base pairs of homology overlap between the parts (page 859, ¶2). Van Leeuwen teaches assembling a plasmid from 8 sections that each had homology to the two adjacent sections (Figure 1).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have applied the method of van Leeuwen to assemble the vector comprising the T7 promoter fused to the guide RNA and express the guide RNA in yeast because it would have amounted to a simple combination of known elements and methods to yield predictable results. One would have a reasonable and predictable expectation of success because Van Leeuwen teaches that any plasmid can be assembled using homologous recombination of parts to complete a plasmid using the method in yeast. One would have been motivated to apply the method to adding the targeting sequence of the guide RNA to a plasmid containing the structural components of the guide RNA and the T7 promoter so that the targeting sequence of the guide RNA can be easily changed to target different loci.
Claim 14 is rejected under 35 U.S.C. 103 as being unpatentable over DiCarlo (DiCarlo et al., Nucleic Acids Research (2013), 41: 4336-4343; of record), Zhang II (US 20140357530 A1, published December 4, 2014; of record), Wagner (Wagner et al., Nature Methods (2014), 11: 915-918; of record) and Benton (Benton et al., Molecular and Cellular Biology (1990), 10: 353-360; of record) as applied to claims 1-4, 6, 8-10, 12-13, 15-18 and 20-26 above, and further in view of Horwitz (Horwitz et al., Cell Systems (2015), 1: 88-93; of record) and Gao (Gao et al., DNA Repair (2016), 42: 1-10; of record). This is a maintained rejection.
The teachings of DiCarlo, Zhang II, Wagner and Benton are recited above and applied as for claims 1-4, 6, 8-10, 12-13, 15-18 and 20-26. DiCarlo also teaches that Cas9-mediated double strand breaks can be resolved either through homologous recombination or through error-prone non-homologous end joining (page 4336, ¶2). DiCarlo teaches that Cas9 creates indels at the targeted genomic site (Fig 2D).
DiCarlo, Zhang II, Wagner and Benton do not teach the cells are deficient in an NHEJ component.
Horwitz teaches using CRISPR/Cas9 systems in S. cerevisiae and the yeast K. lactis to integrate DNA constructs into targeted locations (In brief; Abstract). Horwitz teaches deleting KU80 in the K. lactis strain (i.e., making the cell deficient in an NHEJ component) to reduce rates of non-homologous end joining thereby promoting donor insertion by homologous recombination (page 94, ¶2).
Gao teaches that NHEJ pathways of DNA repair are active in G1 and utilize the Ku DNA-end binding protein Ku70/Ku80 (¶ spanning pages 1-2). Gao teaches that knocking out Ku80 does not affect the viability of yeast cells under normal growing conditions (Fig 1). Gao teaches knocking out Ku80 in S. cerevisiae reduces the ability of yeast cells to repair DNA in G1 phase by NHEJ (Fig 3).
It would have been obvious to one skilled in the art before the effective filing date of the claimed invention to have deleted Ku80 from the S. cerevisiae cells in DiCarlo with the T7 promoter driven gRNA rendered obvious above when supplying the yeast with a donor DNA. It would have amounted to the simple combination of known elements by known means to yield predictable results. The skilled artisan would have predicted that Ku80 could be deleted because Gao demonstrates knocking out components of the NHEJ pathway is routine and does not reduce yeast viability. The skilled artisan would have been motivated to do so for the purpose of promoting donor DNA integration and reducing indel formation caused by Cas9-mediated double strand breaks.
Response to Arguments
Applicant argues that the prior art of record does not disclose a working system for expression of guide RNAs in a yeast cell using a viral-subunit DNA-dependent RNA polymerase. Applicant then argues that DiCarlo teaches expression of a guide RNA from an SNR52 promoter, a eukaryotic Pol III promoter, in yeast (Remarks, page 6, ¶2). These arguments have been fully considered but are not persuasive because the §103 rejection is based on the combination of teachings in the cited references. If the prior art of DiCarlo had disclosed a system for expression of guide RNAs in a yeast cell using a viral-subunit DNA-dependent RNA polymerase, then the claims would have been rejected for anticipation under §102. Therefore, Applicant’s argument does not address the merits of the rejection. Additionally, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986).
Applicant argues that although Zhang II teaches that a T7 promoter can drive guide RNA expression, Zhang II does not demonstrate that a T7 promoter can actually function to express guide RNAs in eukaryotic cells. Applicant argues that Zhang II only mentions yeast as a host for amplifying vectors (page 6, ¶3). These arguments have been fully considered but are not persuasive. First, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. Second, MPEP 2143.02 teaches that obviousness does not require absolute predictability, but only a reasonable expectation of success is required. Because Wagner demonstrates guide RNA expression in a different eukaryotic cell and Benton demonstrates that the T7 promoter and T7 RNAP system is functional in S. cerevisiae (i.e., a yeast), the skilled artisan would have a reasonable expectation that the T7 promoter/RNAP system could be used in DiCarlo’s S. cerevisiae to drive guide RNA expression. Finally, Zhang II teaches that generically the T7 promoter/RNAP can be used in eukaryotic cells. S. cerevisiae is a well-known eukaryotic cell model that is used to study fundamental eukaryotic cell biology. As such, the skilled artisan would have understood Zhang II’s statement “to express guide RNAs in eukaryotic cells, Applicants express the T7 polymerase in eukaryotic cells to drive expression of guide RNAs using the T7 promoter” ([0468]) could reasonably include S. cerevisiae cells.
Applicant argues that Wagner would not have provided the skilled artisan a reasonable expectation that the T7 promoter/RNAP would work in yeast because results from one eukaryote cannot simply be extrapolated to another eukaryote. Applicant argues that this is because the U6 promoter that works in yeast does not work in Plasmodium (page 7, ¶2). This argument has been fully considered but is not persuasive. First, Wagner does not teach that the U6 promoter is non-functional in Plasmodium. Wagner instead states that the “[U6] promoter has not been well defined in P. falciparum.”, and because of this, the U6 promoter was not chosen for developing a guide RNA expression system in Plasmodium (page 915, ¶2). It is unknown whether, once defined, a U6 Pol III promoter could be used. Second, the question at hand is not whether the U6 promoter can be used in Plasmodium, but whether there was a reasonable expectation that that T7 promoter/RNAP would be functional in yeast. Benton provides the evidence that T7 promoter/RNAP system is functional for RNA production in S. cerevisiae.
Applicant argues that the RNAs produced by the T7 promoter/RNAP system in yeast by Benton were not functional (page 7, ¶3). This argument has been fully considered but is not persuasive because the RNAs tested by Benton were protein-encoding mRNAs. CRISPR guide RNAs are noncoding RNAs and do not need to be capped or interact with the translational machinery. As such, for a reasonable expectation of success the skilled artisan need only determine whether the guide RNA would be produced in yeast by the T7 promoter/RNAP. Because Benton teaches two different RNAs can be produced using the T7 promoter/RNAP system in S. cerevisiae, the skilled artisan would predict that DiCarlo’s guide RNA would also be produced when cloned under the control of the T7 promoter.
Applicant argues that Enayati teaches T7 RNAP codon optimization in Aspergillus, which is not a yeast, and for using the T7 system for expressing an RNA interfering noncoding RNA (RNAi). Applicant argues that this system would not provide a reasonable expectation of success because 1) S. cerevisiae lacks an RNAi pathway and 2) the T7-generated transcripts in S. cerevisiae would likely be degraded by the yeast nuclear exosome system instead of generating precise CRSPR guide RNAs (Remarks, page 8, last ¶). These arguments have been fully considered but are not persuasive. First, it does not address the merits of the rejection. Enayati was relied upon for teaching that codon-optimization in fungal cells, including the gene encoding T7 RNAP, was known in the art, and not for the reasonable expectation of guide RNA expression from the T7 promoter/RNAP system, which was already rendered obvious for claim 1. Second, guide RNAs are non-coding RNAs and do not need to be capped. So, the fact that the T7-produced transcripts are uncapped would not have dissuaded the skilled artisan from using the T7 promoter to drive expression. Third, Applicant does not provide evidence that unprocessed noncoding RNAs are degraded in yeast. MPEP 716.01(c) makes clear that arguments of counsel cannot take the place of evidence in the record. Furthermore, Wagner teaches the T7 RNAP produces RNAs of “defined lengths” when used with the T7 terminator. As such, the skilled artisan would not have predicted that the guide RNAs would not have “ragged” 3’ ends. In any event though, Cas9 guide RNAs were known in the prior art to be amenable to tolerating additional nucleotides appended to their 3’ ends. See e.g., Mali et al., Nature Methods (2013), 10: 957-963. As such, the skilled artisan would have predicted that even if additional nucleotides were included on the 3’ end of the guide RNAs, they would still be able to bind to Cas9 and direct Cas9-mediated genome editing.
Applicant argues that Jones, van Leeuwen, Horvitz and/or Gao do not remedy the deficiencies of DiCarlo, Zhang II, Wagner and Benton (Remarks, pages 9-11). These arguments have been fully considered but are not persuasive because a prima facie case of obviousness is established for claim 1 for the reasons set for in the §103 rejections of record and the response to Applicant’s arguments in paragraphs 50-53 above.
Conclusion
No claims are allowable.
THIS ACTION IS MADE FINAL. 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 nonprovisional extension fee (37 CFR 1.17(a)) 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 mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to CATHERINE KONOPKA whose telephone number is (571)272-0330. The examiner can normally be reached Mon - Fri 7- 4.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Ram Shukla can be reached at (571)272-0735. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/CATHERINE KONOPKA/Examiner, Art Unit 1635