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 .
Applicant’s election without traverse of Group I in the reply filed on June 8, 2026 is acknowledged. Additionally in response to the election of species requirements the Applicants elected the following without traverse: (i) the Cas nuclease being LbCas12a; (ii) the sequence engineering changes the activity of the Cas nuclease in interacting with a PAM sequence.
Claims 1-7, 11, 14, 16-17, 21-23, 28-31, 45, and 48 are currently pending.
Claims 5, 22-23, 28-29, 45 and 48 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to nonelected subject matter, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on June 8, 2026.
Summary of Requirements for Patent Applications Filed On Or After July 1, 2022, That Have Sequence Disclosures
3. 37 CFR 1.831(a) requires that patent applications which contain disclosures of nucleotide and/or amino acid sequences that fall within the definitions of 37 CFR 1.831(b) must contain a “Sequence Listing XML”, as a separate part of the disclosure, which presents the nucleotide and/or amino acid sequences and associated information using the symbols and format in accordance with the requirements of 37 CFR 1.831-1.835. This “Sequence Listing XML” part of the disclosure may be submitted:
1. In accordance with 37 CFR 1.831(a) using the symbols and format requirements of 37 CFR 1.832 through 1.834 via the USPTO patent electronic filing system (see Section I.1 of the Legal Framework for Patent Electronic System (https://www.uspto.gov/PatentLegalFramework), hereinafter “Legal Framework”) in XML format, together with an incorporation by reference statement of the material in the XML file in a separate paragraph of the specification (an incorporation by reference paragraph) as required by 37 CFR 1.835(a)(2) or 1.835(b)(2) identifying:
a. the name of the XML file
b. the date of creation; and
c. the size of the XML file in bytes; or
2. In accordance with 37 CFR 1.831(a) using the symbols and format requirements of 37 CFR 1.832 through 1.834 on read-only optical disc(s) as permitted by 37 CFR 1.52(e)(1)(ii), labeled according to 37 CFR 1.52(e)(5), with an incorporation by reference statement of the material in the XML format according to 37 CFR 1.52(e)(8) and 37 CFR 1.835(a)(2) or 1.835(b)(2) in a separate paragraph of the specification identifying:
a. the name of the XML file;
b. the date of creation; and
c. the size of the XML file in bytes.
SPECIFIC DEFICIENCIES AND THE REQUIRED RESPONSE TO THIS NOTICE ARE AS FOLLOWS:
Specific deficiency - Sequences appearing in the drawings are not identified by sequence identifiers in accordance with 37 CFR 1.831(c). Sequence identifiers for sequences (i.e., “SEQ ID NO:X” or the like) must appear either in the drawings or in the Brief Description of the Drawings. See Fig 25 A.
Required response – Applicant must provide:
Amended drawings in accordance with 37 CFR 1.121(d) inserting the required sequence identifiers;
AND/OR
A substitute specification in compliance with 37 CFR 1.52, 1.121(b)(3), and 1.125 inserting the required sequence identifiers (i.e., “SEQ ID NO:X” or the like) into the Brief Description of the Drawings, consisting of:
• A copy of the previously-submitted specification, with deletions shown with strikethrough or brackets and insertions shown with underlining (marked-up version);
• A copy of the amended specification without markings (clean version); and
• A statement that the substitute specification contains no new matter.
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 7, 11, and 16-17 are 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 7 is rejected over the recitation of the phrase “wherein the modification is with a group”. This recitation is indefinite because it is unclear what it means for a modification to be with a group.
Claim 11 is rejected over the recitation of the phrase “5’/3’ DNA-/RNA-extended CRISPER RNA”. This recitation is indefinite because it is unclear what the “/”. For example does it mean that the guide is truncated at either or both ends? Further it’s unclear if the guide comprises DNA, RNA, or both. Clarification is requested.
Claims 16-17 are rejected over the recitation of the phrase “the nucleotides” in claims 16 and 17. There is insufficient antecedent basis for this limitation in the claim.
Claim 17 is rejected over the recitation of the phrase “the modification is with a group”. There is insufficient antecedent basis for this limitation in the claim. Additionally it is unclear what it means for a modification to be with a group. Finally it is not clear what is being modified. Clarification is required.
Claim Rejections - 35 USC § 102
5. The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 1 and 14 are rejected under 35 U.S.C. 102(a)(2) as being anticipated by Gootenberg (WO 2021/163584 Filed 2/12/2021 with priority to 63/075,684 Filed 9/8/2020).
Regarding Claim 1 Gootenberg discloses a one-pot LAMP-Casl2b reaction to detect SARS-CoV-2. Gootenberg teaches that the reactions were performed using IX Isothermal Amplification Buffer, 1.4 mM dNTPs, 6.4 units of Bst2.0 WarmStart DNA Polymerase, 6 units of WarmStart RTx Reverse Transcriptase, 250 nM Casl2b protein, 250 nM sgRNA, 200 nM fluorescent reporter, 1.6 µM FIP/BIP primers, 0.2 µM F3/B3 primers, 0.4 µM LoopF/B primers and variable concentrations of MgSO4 20 µL reactions. Where indicated, mock clinical sample with SARS-CoV-2 genomic standard was added at 10% of the final reaction volume to simulate crude input from sample lysis. One-pot reactions were performed at 60 °C (unless otherwise indicated) for 1-2 hrs on a qPCR machine (BioRad) with fluorescent measurements every 2 minutes (para 0559). Gootenberg further teaches that the guide polynucleotide comprises a spacer specific for the N gene or S gene of SARS-CoV-2 (para 0009). Thus Gootenberg teaches a method for detecting a target polynucleotide, comprising incubating the target polynucleotide in a mixture that comprises (a) a polymerase, (b) deoxynucleoside triphosphates (dNTPs), (c) primers for amplifying the target polynucleotide, (c) a CRISPR- associated (Cas) nuclease, and (d) a guide RNA comprising a spacer fragment complementary to a target fragment on the target polynucleotide, under conditions so that the polymerase effectively amplifies the target polynucleotide while the Cas nuclease is capable of cleaving the amplified target polynucleotide.
Regarding Claim 14 Gootenberg discloses a tracr (trans-activating CRISPR) sequence (e.g. tracrRNA or an active partial tracrRNA) (para 0135). Thus Gootenberg teaches a method wherein the guide RNA includes a truncated trans-activating crispr RNA (tracrRNA) sequence.
Claim Rejections - 35 USC § 103
6. 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.
7. Claims 2-4, 21, and 30-31 are rejected under 35 U.S.C. 103 as being unpatentable over Gootenberg (WO 2021/163584 Filed 2/12/2021 with priority to 63/075,684 Filed 9/8/2020) in view of Toth (Nucleic Acids Research Feb 28, 2020 Vol 48 No 7).
The teachings of Gootenberg are presented above.
Gootenberg does not teach a method wherein the target fragment for the guide RNA includes or is adjacent to, a protospacer adjacent motif (PAM) sequence recognizable by the Cas nuclease, which PAM sequence is suboptimal (clm 2). Gootenberg does not teach a method wherein the PAM sequence is not canonical (clm 3). Gootenberg does not teach a method wherein the Cas nuclease is LbCas12a, and the PAM sequence is selected from the group consisting of NTTV, TNTV, TTNV, TTNT, VTTT, TVTT, VVTT, VTVT, VNVV, NVNV, NVVV, VNTV, NTVV, TNVV, YYYN and VVNV, wherein N denotes any nucleotide (clm 4).
However Toth teaches that LbCas12a wild type shows robust activity on its optimal PAM sequences (TTTV, where V is A, C, or G) and lower but considerable activity on TTTT (page 3725, col 2 Fig 2). In the instant case the TTTT PAM sequence is considered to be suboptimal since it has weaker activity. Additionally the TTTT PAM sequence is considered not canonical since it is not TTTV. Finally the TTTT PAM sequence has the TTNT PAM sequence, wherein N is any nucleotide.
Accordingly, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Gootenberg by using a Cas nuclease that recognizes a suboptimal/non-canonical PAM sequence as suggested by Toth. Based on the teaching of Toth the skilled artisan would have been motivated to use a Cas nuclease (such as LbCas12a) that recognizes a suboptimal/non-canonical PAM sequence (such as TTTT) for the benefit of being able to cut at both canonical and non-canonical sites.
Gootenberg does not teach a method wherein the Cas nuclease sequence in engineered (clm 21). Gootenberg does not teach a method wherein the sequence engineering changes the activity of the Cas nuclease interacting with a PAM sequence (clm 30). Gootenberg does not teach a method wherein the LbCas12a has substitution at Lys538 or Tyr542 (clm 31).
However Toth teaches that they made LbCas12a variants with altered PAM specificities to further broaden the genome targeting range of Cas12a nucleases (page 3722). Toth teaches that they created an improved LbCas12a variant (termed impLbCas12a, possessing overall the mutations D156R, G532R, K538V, Y542R and K595R).
Accordingly, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Gootenberg by engineering variant Cas nucleases that change the activity of the Cas nuclease interacting with a PAM sequence as suggested by Toth. One of skill in the art would have been motivated to engineer variant Cas nucleases that change the activity of the Cas nuclease interacting with a PAM sequence for the benefit of broadening the genome targeting range. Further one of skill in the art would have been motivated to make a LbCas12a mutant with substitutions at Lys538 or Tyr542 since Toth discloses a mutant having these mutations and teaches that they provide increased efficiency for genome editing and transcriptome modulating applications (abstract).
8. Claims 6-7 are rejected under 35 U.S.C. 103 as being unpatentable over Gootenberg (WO 2021/163584 Filed 2/12/2021 with priority to 63/075,684 Filed 9/8/2020) in view of Chen (US 2018/0335424 Pub 11/22/2018).
The teachings of Gootenberg are presented above.
Gootenberg does not teach a method wherein at least one of the dNTPs is modified (clm 6). Gootenberg does not teach a method wherein the modification is selected from the group consisting of phosphoryl, biotin, digoxigenin, amino, thiol, phosphorthioate, and methyl (clm 7).
However Chen discloses that labeled amplified DNA fragments can be generated by incorporation of one or more modified dNTPs that are labeled with an affinity-binding molecule or a detectable moiety during the amplification reaction. Chen teaches incorporation of one or more modified dNTPs that has an amino allyl-group, a biotin group, a fluorescent or other detectable dye, or another moiety that permits it to be detected, either directly, or indirectly following labeling with any other detectable molecule or combination of molecules known in the art. (para 0323).
Accordingly, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Gootenberg by using dNTPs modified with biotin as suggested by Chen. One of skill in the art would have been motivated to use dNTPs modified with biotin for the benefit of being able to add a detectable label to the amplification products.
9. Claim 11 is rejected under 35 U.S.C. 103 as being unpatentable over Gootenberg (WO 2021/163584 Filed 2/12/2021 with priority to 63/075,684 Filed 9/8/2020) in view of Cheng (US 2019/0002889 Pub 1/3/2019).
The teachings of Gootenberg are presented above.
Gootenberg does not teach a method wherein the guide RNA comprises a truncated or 5'/3' DNA-/RNA-extended CRISPR RNA (crRNA) that includes the spacer fragment.
However Cheng discloses a crRNA includes a truncated direct repeat sequence and a spacer sequence (para 0218).
Accordingly, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Gootenberg by using a crRNA comprising a truncated direct repeat sequence and a spacer sequence as suggested by Cheng. One of skill in the art would have been motivated to use a crRNA comprising a truncated direct repeat sequence and a spacer sequence for the benefit of minimizing off target cleavage.
10. Claims 16-17 are rejected under 35 U.S.C. 103 as being unpatentable over Gootenberg (WO 2021/163584 Filed 2/12/2021 with priority to 63/075,684 Filed 9/8/2020) in view of Fox (US 2020/0263189 Pub 8/20/2020).
The teachings of Gootenberg are presented above.
Gootenberg does not teach a method wherein at least one of the nucleotides in the spacer fragment is a not a standard ribonucleotide (clm 16). Gootenberg does not teach a method wherein at least one of the nucleotides in the spacer fragment is selected from the group consisting of a deoxynucleotide, a locked nucleic acid (LNA), a bridged nucleic acid (BNA), a deoxy uridine, a deoxy inosine, a pseudouridin, a methylpseudouridin, and modified nucleotide, wherein the modification is with a group selected from the group consisting of phosphoryl, biotin, digoxigenin, amino, thiol, phosphorthioate, methyl, 2'-O-methyl- 3'-phosphonoacetate (MP), 2'O-methoxyethyl (MOE), Fluoro(F), S-constrained ethyl, 2'-0-methyl- PS (MS) and 2'-O-methyl-thioPACE (MSP) (clm 17).
However Fox teaches that a guide polynucleotide can comprise at least one nucleotide, phosphodiester bond or linkage modification such as, but not limited, to Locked Nucleic Acid (LNA), 5-methyl dC, 2,6-Diaminopurine, 2′-Fluoro A, 2′-Fluoro U, 2′-O-Methyl RNA, phosphorothioate bond, linkage to a cholesterol molecule, linkage to a polyethylene glycol molecule, linkage to a spacer 18 (hexaethylene glycol chain) molecule, or 5′ to 3′ covalent linkage resulting in circularization (0177).
Accordingly, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of Gootenberg by using a guide RNA with a modified spacer (such as LNA) as suggested by Fox. One of skill in the art would have been motivated to use a guide RNA with a modified spacer (such as LNA) for the benefit of increasing target specificity.
11. Any inquiry concerning this communication or earlier communications from the examiner should be directed to AMANDA HANEY whose telephone number is (571)272-8668. The examiner can normally be reached Monday-Friday, 8:15am-4:45pm EST.
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, Wu-Cheng Shen can be reached at 571-272-3157. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/AMANDA HANEY/Primary Examiner, Art Unit 1682
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