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
This action is written in response to applicant’s correspondence received 02/12/2024. Claims 1-15 are currently pending and are examined herein.
Objections to the Specification
The use of the terms “Wizard® SV Gel and PCR Clean-Up System”, “NEBuilder® HiFi DNA Assembly Master Mix”, and “GeneJET™ Plasmid Miniprep Kit”, which are trade names or marks used in commerce, has been noted in this application. Please see p. 73. The terms should be accompanied by the generic terminology; furthermore the terms should be capitalized wherever they appear or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM , or ® following the term.
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) are permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Claim Rejections - 35 USC § 112(b)
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 13-14 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.
Claims 13-14 all recite the limitation "”the non-adapting CRISPR-Cas system” There is insufficient antecedent basis for this limitation in the claim. Claim 11, from which the instantly rejected claims depend, does not recite a non-adapting CRISPR-Cas system.
Claim Rejections - 35 USC § 101
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Claims 1-14 are rejected under 35 U.S.C. 101 because the claimed invention is directed to product of nature, or natural phenomenon, without significantly more.
Claim interpretation: The claims recite Cas1 and Cas2 polypeptides. The broadest reasonable interpretation of the terms encompasses both full length, canonical, functional Cas1/Cas2 polypeptides, and truncated, mutated, and/or nonfunctional isoforms.
Subject Matter Eligibility Analysis:
Claims 1, 3, 4 and 5:
Step 1: Is the claim to a process, machine, manufacture or composition of matter? (MPEP 2106.03.II)
Claim 1 recites a polynucleotide comprising a nucleic acid sequence encoding a CRISPR Cas1 polypeptide. The claim is drawn to a composition of matter and is eligible at step 1. Claims 4 and 5 specify that the Cas1 polypeptide is a polypeptide of a Lactococcus bacterial strain, or that it comprises an amino acid sequence set forth in SEQ ID NO: 2, which are also compositions of matter.
Step 2A Prong 1: Does the claim recite an abstract idea, law of nature, product of nature or natural phenomenon? (MPEP 2106.04.II.A.1) If the claim recites a product of nature, is the product of nature markedly different from its closest naturally occurring counterpart? (MPEP 2106.04(c))
Claim 1 recites a polynucleotide encoding a Cas1 polypeptide. This refers to a judicial exception which is a product of nature or natural phenomenon, as evidenced by UniProt Reference Sequence A0A6H0UGI6 (submitted Dec 2019), which is a naturally-occurring Cas1 amino acid sequence from Lactococcus raffinolactis with 100% identity to SEQ ID NO: 2 and the closest naturally-occurring counterpart to that specific cas1:
RESULT 1
A0A6H0UGI6_9LACT
ID A0A6H0UGI6_9LACT Unreviewed; 302 AA.
AC A0A6H0UGI6;
DT 12-AUG-2020, integrated into UniProtKB/TrEMBL.
DT 12-AUG-2020, sequence version 1.
DT 08-OCT-2025, entry version 15.
DE RecName: Full=CRISPR-associated endonuclease Cas1 {ECO:0000256|HAMAP-Rule:MF_01470};
DE EC=3.1.-.- {ECO:0000256|HAMAP-Rule:MF_01470};
GN Name=cas1 {ECO:0000256|HAMAP-Rule:MF_01470,
GN ECO:0000313|EMBL:QIW54544.1};
GN ORFNames=GU336_10590 {ECO:0000313|EMBL:QIW54544.1};
OS Pseudolactococcus raffinolactis.
OC Bacteria; Bacillati; Bacillota; Bacilli; Lactobacillales; Streptococcaceae;
OC Pseudolactococcus.
OX NCBI_TaxID=1366 {ECO:0000313|EMBL:QIW54544.1, ECO:0000313|Proteomes:UP000501945};
RN [1] {ECO:0000313|EMBL:QIW54544.1, ECO:0000313|Proteomes:UP000501945}
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=Lr_19_5 {ECO:0000313|EMBL:QIW54544.1,
RC ECO:0000313|Proteomes:UP000501945};
RA Ybazeta G., Ross M., Brabant-Kirwan D., Saleh M., Dillon J.A., Splinter K.,
RA Nokhbeh R.;
RT "Whole genome sequences of Lactococcus raffinolactis strains isolated from
RT sewage.";
RL Submitted (DEC-2019) to the EMBL/GenBank/DDBJ databases.
CC -!- FUNCTION: CRISPR (clustered regularly interspaced short palindromic
CC repeat), is an adaptive immune system that provides protection against
CC mobile genetic elements (viruses, transposable elements and conjugative
CC plasmids). CRISPR clusters contain spacers, sequences complementary to
CC antecedent mobile elements, and target invading nucleic acids. CRISPR
CC clusters are transcribed and processed into CRISPR RNA (crRNA). Acts as
CC a dsDNA endonuclease. Involved in the integration of spacer DNA into
CC the CRISPR cassette. {ECO:0000256|HAMAP-Rule:MF_01470}.
CC -!- COFACTOR:
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Evidence={ECO:0000256|HAMAP-Rule:MF_01470};
CC Name=Mn(2+); Xref=ChEBI:CHEBI:29035;
CC Evidence={ECO:0000256|HAMAP-Rule:MF_01470};
CC -!- SUBUNIT: Homodimer, forms a heterotetramer with a Cas2 homodimer.
CC {ECO:0000256|ARBA:ARBA00038592, ECO:0000256|HAMAP-Rule:MF_01470}.
CC -!- SIMILARITY: Belongs to the CRISPR-associated endonuclease Cas1 family.
CC {ECO:0000256|HAMAP-Rule:MF_01470}.
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DR EMBL; CP047616; QIW54544.1; -; Genomic_DNA.
DR RefSeq; WP_167839034.1; NZ_CP047616.1.
DR AlphaFoldDB; A0A6H0UGI6; -.
DR Proteomes; UP000501945; Chromosome.
DR GO; GO:0003677; F:DNA binding; IEA:UniProtKB-KW.
DR GO; GO:0004520; F:DNA endonuclease activity; IEA:InterPro.
DR GO; GO:0016787; F:hydrolase activity; IEA:UniProtKB-KW.
DR GO; GO:0046872; F:metal ion binding; IEA:UniProtKB-UniRule.
DR GO; GO:0051607; P:defense response to virus; IEA:UniProtKB-UniRule.
DR GO; GO:0043571; P:maintenance of CRISPR repeat elements; IEA:UniProtKB-UniRule.
DR Gene3D; 1.20.120.920; CRISPR-associated endonuclease Cas1, C-terminal domain; 1.
DR HAMAP; MF_01470; Cas1; 1.
DR InterPro; IPR050646; Cas1.
DR InterPro; IPR002729; CRISPR-assoc_Cas1.
DR InterPro; IPR042206; CRISPR-assoc_Cas1_C.
DR InterPro; IPR019855; CRISPR-assoc_Cas1_NMENI.
DR NCBIfam; TIGR00287; cas1; 1.
DR NCBIfam; TIGR03639; cas1_NMENI; 1.
DR PANTHER; PTHR34353; CRISPR-ASSOCIATED ENDONUCLEASE CAS1 1; 1.
DR PANTHER; PTHR34353:SF2; CRISPR-ASSOCIATED ENDONUCLEASE CAS1 1; 1.
DR Pfam; PF01867; Cas_Cas1; 1.
PE 3: Inferred from homology;
KW Antiviral defense {ECO:0000256|ARBA:ARBA00023118, ECO:0000256|HAMAP-
KW Rule:MF_01470};
KW DNA-binding {ECO:0000256|ARBA:ARBA00023125, ECO:0000256|HAMAP-
KW Rule:MF_01470};
KW Endonuclease {ECO:0000256|ARBA:ARBA00022759, ECO:0000256|HAMAP-
KW Rule:MF_01470};
KW Hydrolase {ECO:0000256|ARBA:ARBA00022801, ECO:0000256|HAMAP-Rule:MF_01470};
KW Magnesium {ECO:0000256|ARBA:ARBA00022842, ECO:0000256|HAMAP-Rule:MF_01470};
KW Manganese {ECO:0000256|ARBA:ARBA00023211, ECO:0000256|HAMAP-Rule:MF_01470};
KW Metal-binding {ECO:0000256|ARBA:ARBA00022723, ECO:0000256|HAMAP-
KW Rule:MF_01470};
KW Nuclease {ECO:0000256|ARBA:ARBA00022722, ECO:0000256|HAMAP-Rule:MF_01470}.
FT BINDING 150
FT /ligand="Mn(2+)"
FT /ligand_id="ChEBI:CHEBI:29035"
FT /evidence="ECO:0000256|HAMAP-Rule:MF_01470"
FT BINDING 207
FT /ligand="Mn(2+)"
FT /ligand_id="ChEBI:CHEBI:29035"
FT /evidence="ECO:0000256|HAMAP-Rule:MF_01470"
FT BINDING 222
FT /ligand="Mn(2+)"
FT /ligand_id="ChEBI:CHEBI:29035"
FT /evidence="ECO:0000256|HAMAP-Rule:MF_01470"
SQ SEQUENCE 302 AA; 34688 MW; DDE555FF6978A010 CRC64;
Query Match 100.0%; Score 1540; Length 302;
Best Local Similarity 100.0%;
Matches 302; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 1 MRDIIYVENRYFISSRENALRFHDYINKATHFIAFDDIDILVLDNARSYLSNGVINDCLD 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 MRDIIYVENRYFISSRENALRFHDYINKATHFIAFDDIDILVLDNARSYLSNGVINDCLD 60
Qy 61 RNILILTCDNKHSPKAILSNAFANKKRLERLRNQLQLSSKSKNRLWRKIVMAKINNQADA 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 RNILILTCDNKHSPKAILSNAFANKKRLERLRNQLQLSSKSKNRLWRKIVMAKINNQADA 120
Qy 121 VTFTVQDGTVHREIIELGKMVTEGDKDNREAVVARKYFRTLFGGNFKRGRFDDVINSALN 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 VTFTVQDGTVHREIIELGKMVTEGDKDNREAVVARKYFRTLFGGNFKRGRFDDVINSALN 180
Qy 181 YGYALVRAVIRRELAICGFEMSFGIHHMSTENPFNLSDDMIEVFRPFVDVLVFEIIVTND 240
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 YGYALVRAVIRRELAICGFEMSFGIHHMSTENPFNLSDDMIEVFRPFVDVLVFEIIVTND 240
Qy 241 VTVFDYEIKKLLVNIFLEKCVIDGKVMSLTDAVRVTIQSLITCLEDDSSAPLKLPSFIQE 300
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 241 VTVFDYEIKKLLVNIFLEKCVIDGKVMSLTDAVRVTIQSLITCLEDDSSAPLKLPSFIQE 300
Qy 301 GK 302
||
Db 301 GK 302
Because the claim is drawn to a product of nature, it is subject to the markedly different characteristics analysis, as described above. In the instant case, the closest naturally occurring counterpart is the L. raffinolactis Cas1 amino acid sequence and the gene encoding it, as seen above. This is further supported by FIG. 1 of the instant application, which shows the structure of naturally-occurring CRISPR-Cas operons for L. raffinolactis and L. cremoris. There is no structural or functional difference between the naturally-occurring amino acid sequence and the claimed sequence. The same logic applies to claims 3,
Please note that while the above listing refers to Pseudolactococcus raffinolactis, P. raffinolactis was recently reclassified from Lactococcus and was formerly known as L. raffinolactis, as disclosed by Cha (see Cha et al. Complete genome sequence of Pseudolactococcus raffinolactis strain GCULR from a spotted seal (Phoca largha) in Korea. BMC Genomic Data (2025) 26:93.)(Abstract).
It is also relevant to note that cas1 and cas2 genes are, “notably conserved across the three major types of CRISPR systems” and are present in a variety of prokaryotes, including E. coli (Nuñez et al. Cas1–Cas2 complex formation mediates spacer acquisition during CRISPR–Cas adaptive immunity. Nature Structural & Molecular Biology volume 21, pages528–534 (2014))(p. 528 2nd ¶). Therefore, polynucleotides (genes and genomes) encoding cas1 polypeptides or cas1 and cas2 polypeptides are widespread among CRISPR/Cas systems and among other prokaryotes, and there are no additional structural or functional characteristics recited by claims 1, 3, 4 or 5 which render the generic cas1/cas2 sequences markedly different from their naturally occurring counterparts, i.e., genomes or plasmids encoding cas1 and/or cas2 polypeptides.
Step 2A Prong 2: Does the claim recite additional elements that integrate the judicial exception into a practical application? (MPEP 2106.04.II.A.2, MPEP 2106.04(d))
Step 2B: Do the additional elements amount to significantly more than the judicial exception? (MPEP 2106.05)
Regarding steps 2A prong two and 2B, the claims do not recite any additional elements, and therefore do not recite additional elements that integrate the judicial exception into a practical application or amount to significantly more than the judicial exception.
Regarding claims 2, 6 and 7, which recite the polynucleotide comprising a Cas1 and Cas2 sequence (claims 2, 6, 7) from a Lactococcus (claim 6) and/or comprising an amino acid sequence having at least 85% identity to SEQ ID NO: 4 (claim 7). In addition to the cas1 gene discussed above, L. raffinolactis also naturally comprises a cas2 gene (UniProt A0A6H0UID8, submitted Dec. 2019) encoding an amino acid sequence with 100% identity to SEQ ID NO: 4:
RESULT 1
A0A6H0UID8_9LACT
ID A0A6H0UID8_9LACT Unreviewed; 101 AA.
AC A0A6H0UID8;
DT 12-AUG-2020, integrated into UniProtKB/TrEMBL.
DT 12-AUG-2020, sequence version 1.
DT 08-OCT-2025, entry version 12.
DE RecName: Full=CRISPR-associated endoribonuclease Cas2 {ECO:0000256|HAMAP-Rule:MF_01471};
DE EC=3.1.-.- {ECO:0000256|HAMAP-Rule:MF_01471};
GN Name=cas2 {ECO:0000256|HAMAP-Rule:MF_01471,
GN ECO:0000313|EMBL:QIW54543.1};
GN ORFNames=GU336_10585 {ECO:0000313|EMBL:QIW54543.1};
OS Pseudolactococcus raffinolactis.
OC Bacteria; Bacillati; Bacillota; Bacilli; Lactobacillales; Streptococcaceae;
OC Pseudolactococcus.
OX NCBI_TaxID=1366 {ECO:0000313|EMBL:QIW54543.1, ECO:0000313|Proteomes:UP000501945};
RN [1] {ECO:0000313|EMBL:QIW54543.1, ECO:0000313|Proteomes:UP000501945}
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=Lr_19_5 {ECO:0000313|EMBL:QIW54543.1,
RC ECO:0000313|Proteomes:UP000501945};
RA Ybazeta G., Ross M., Brabant-Kirwan D., Saleh M., Dillon J.A., Splinter K.,
RA Nokhbeh R.;
RT "Whole genome sequences of Lactococcus raffinolactis strains isolated from
RT sewage.";
RL Submitted (DEC-2019) to the EMBL/GenBank/DDBJ databases.
CC -!- FUNCTION: CRISPR (clustered regularly interspaced short palindromic
CC repeat), is an adaptive immune system that provides protection against
CC mobile genetic elements (viruses, transposable elements and conjugative
CC plasmids). CRISPR clusters contain sequences complementary to
CC antecedent mobile elements and target invading nucleic acids. CRISPR
CC clusters are transcribed and processed into CRISPR RNA (crRNA).
CC Functions as a ssRNA-specific endoribonuclease. Involved in the
CC integration of spacer DNA into the CRISPR cassette. {ECO:0000256|HAMAP-
CC Rule:MF_01471}.
CC -!- COFACTOR:
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Evidence={ECO:0000256|ARBA:ARBA00001946,
CC ECO:0000256|HAMAP-Rule:MF_01471};
CC -!- SUBUNIT: Homodimer, forms a heterotetramer with a Cas1 homodimer.
CC {ECO:0000256|HAMAP-Rule:MF_01471}.
CC -!- SIMILARITY: Belongs to the CRISPR-associated endoribonuclease Cas2
CC protein family. {ECO:0000256|ARBA:ARBA00009959, ECO:0000256|HAMAP-
CC Rule:MF_01471}.
CC ---------------------------------------------------------------------------
CC Copyrighted by the UniProt Consortium, see https://www.uniprot.org/terms
CC Distributed under the Creative Commons Attribution (CC BY 4.0) License
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DR EMBL; CP047616; QIW54543.1; -; Genomic_DNA.
DR RefSeq; WP_167839033.1; NZ_CP047616.1.
DR AlphaFoldDB; A0A6H0UID8; -.
DR Proteomes; UP000501945; Chromosome.
DR GO; GO:0016787; F:hydrolase activity; IEA:UniProtKB-KW.
DR GO; GO:0046872; F:metal ion binding; IEA:UniProtKB-UniRule.
DR GO; GO:0004521; F:RNA endonuclease activity; IEA:InterPro.
DR GO; GO:0051607; P:defense response to virus; IEA:UniProtKB-UniRule.
DR GO; GO:0043571; P:maintenance of CRISPR repeat elements; IEA:UniProtKB-UniRule.
DR HAMAP; MF_01471; Cas2; 1.
DR InterPro; IPR021127; CRISPR_associated_Cas2.
DR InterPro; IPR019199; Virulence_VapD/CRISPR_Cas2.
DR NCBIfam; TIGR01573; cas2; 1.
DR Pfam; PF09827; CRISPR_Cas2; 1.
DR SUPFAM; SSF143430; TTP0101/SSO1404-like; 1.
PE 3: Inferred from homology;
KW Antiviral defense {ECO:0000256|ARBA:ARBA00023118, ECO:0000256|HAMAP-
KW Rule:MF_01471};
KW Endonuclease {ECO:0000256|ARBA:ARBA00022759, ECO:0000256|HAMAP-
KW Rule:MF_01471};
KW Hydrolase {ECO:0000256|ARBA:ARBA00022801, ECO:0000256|HAMAP-Rule:MF_01471};
KW Magnesium {ECO:0000256|ARBA:ARBA00022842, ECO:0000256|HAMAP-Rule:MF_01471};
KW Metal-binding {ECO:0000256|ARBA:ARBA00022723, ECO:0000256|HAMAP-
KW Rule:MF_01471};
KW Nuclease {ECO:0000256|ARBA:ARBA00022722, ECO:0000256|HAMAP-Rule:MF_01471}.
FT BINDING 8
FT /ligand="Mg(2+)"
FT /ligand_id="ChEBI:CHEBI:18420"
FT /ligand_note="catalytic"
FT /evidence="ECO:0000256|HAMAP-Rule:MF_01471"
SQ SEQUENCE 101 AA; 11834 MW; 30C2FBBA8AD79157 CRC64;
Query Match 100.0%; Score 503; Length 101;
Best Local Similarity 100.0%;
Matches 101; Conservative 0; Mismatches 0; Indels 0; Gaps 0;
Qy 1 MMLLCAFDLPRETKEERKAANKYRKRLVELGFAMKQFSLYEREVRHIDVKNRLIDILREE 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 MMLLCAFDLPRETKEERKAANKYRKRLVELGFAMKQFSLYEREVRHIDVKNRLIDILREE 60
Qy 61 LPDTGAITLYLLPNEVNDAQITILGEKSVNKTVRVARIIFL 101
|||||||||||||||||||||||||||||||||||||||||
Db 61 LPDTGAITLYLLPNEVNDAQITILGEKSVNKTVRVARIIFL 101
Claims 8-10 are drawn to vectors comprising the polynucleotide of claims 1, 2 or 3. Applicant defines the term “vector” as, “any nucleic acid molecule into which another nucleic acid molecule (e.g., nucleic acid sequence encoding a Cas polypeptide) can be inserted and which can be introduced into and optionally replicate within a bacterial strain.” (p. 27 ln 19-21). Since the replication aspect is optional, the basic definition of a vector is a nucleic acid molecule into which any other nucleic acid molecule can be inserted, and which can be introduced into a bacterial strain. Isolated polynucleotides comprising the naturally-occurring gene sequences and encoding the naturally-occurring amino acid sequences, which can be introduced into a bacterial cell, meet that definition. MPEP 2106.04 states, “In Myriad, the Supreme Court made clear that not all changes in characteristics will rise to the level of a marked difference, e.g., the incidental changes resulting from isolation of a gene sequence are not enough to make the isolated gene markedly different.”. As a consequence, the claims encompass polynucleotides which are not structurally or functionally distinct from their naturally occurring counterparts.
Regarding claim 11, the claim is drawn to a bacterial strain comprising the polynucleotide of claim 2. The closest naturally occurring counterpart would be L. raffinolactis comprising , and the claim does not recite further characteristics that would differentiate the claimed bacterium from its naturally occurring counterpart.
Regarding claims 12-13, as noted above, the broadest reasonable interpretation of a cas1 or cas2 polypeptide encompasses non-functional truncated variants. The closest naturally occurring counterparts to the recited bacterial strains comprising a cas1 gene, cas2 gene, and a non-adapting CRISPR-Cas system are disclosed by Sampson (Sampson et al. Degeneration of a CRISPR/Cas system and its regulatory target during the evolution of a pathogen. RNA Biol. 2013 Sep 20;10(10):1618–1622.). Sampson discloses strains of Francisella novicida which “harbor degenerated CRISPR/Cas systems” (Abstract) which “lack the ability to integrate new spacer sequences and therefore to adapt to new target sequences” due to truncated cas1 and/or cas2 polypeptides (pp. 1619-20).
Regarding claim 14, it is noted that L. raffinolactis has both cas1 and cas2 genes, and that its cas1 amino acid sequence has 95.5% overall identity to SEQ ID NO: 24:
RESULT 2
A0A6H0UGI6_9LACT
ID A0A6H0UGI6_9LACT Unreviewed; 302 AA.
AC A0A6H0UGI6;
DT 12-AUG-2020, integrated into UniProtKB/TrEMBL.
DT 12-AUG-2020, sequence version 1.
DT 08-OCT-2025, entry version 15.
DE RecName: Full=CRISPR-associated endonuclease Cas1 {ECO:0000256|HAMAP-Rule:MF_01470};
DE EC=3.1.-.- {ECO:0000256|HAMAP-Rule:MF_01470};
GN Name=cas1 {ECO:0000256|HAMAP-Rule:MF_01470,
GN ECO:0000313|EMBL:QIW54544.1};
GN ORFNames=GU336_10590 {ECO:0000313|EMBL:QIW54544.1};
OS Pseudolactococcus raffinolactis.
OC Bacteria; Bacillati; Bacillota; Bacilli; Lactobacillales; Streptococcaceae;
OC Pseudolactococcus.
OX NCBI_TaxID=1366 {ECO:0000313|EMBL:QIW54544.1, ECO:0000313|Proteomes:UP000501945};
RN [1] {ECO:0000313|EMBL:QIW54544.1, ECO:0000313|Proteomes:UP000501945}
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=Lr_19_5 {ECO:0000313|EMBL:QIW54544.1,
RC ECO:0000313|Proteomes:UP000501945};
RA Ybazeta G., Ross M., Brabant-Kirwan D., Saleh M., Dillon J.A., Splinter K.,
RA Nokhbeh R.;
RT "Whole genome sequences of Lactococcus raffinolactis strains isolated from
RT sewage.";
RL Submitted (DEC-2019) to the EMBL/GenBank/DDBJ databases.
CC -!- FUNCTION: CRISPR (clustered regularly interspaced short palindromic
CC repeat), is an adaptive immune system that provides protection against
CC mobile genetic elements (viruses, transposable elements and conjugative
CC plasmids). CRISPR clusters contain spacers, sequences complementary to
CC antecedent mobile elements, and target invading nucleic acids. CRISPR
CC clusters are transcribed and processed into CRISPR RNA (crRNA). Acts as
CC a dsDNA endonuclease. Involved in the integration of spacer DNA into
CC the CRISPR cassette. {ECO:0000256|HAMAP-Rule:MF_01470}.
CC -!- COFACTOR:
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Evidence={ECO:0000256|HAMAP-Rule:MF_01470};
CC Name=Mn(2+); Xref=ChEBI:CHEBI:29035;
CC Evidence={ECO:0000256|HAMAP-Rule:MF_01470};
CC -!- SUBUNIT: Homodimer, forms a heterotetramer with a Cas2 homodimer.
CC {ECO:0000256|ARBA:ARBA00038592, ECO:0000256|HAMAP-Rule:MF_01470}.
CC -!- SIMILARITY: Belongs to the CRISPR-associated endonuclease Cas1 family.
CC {ECO:0000256|HAMAP-Rule:MF_01470}.
CC ---------------------------------------------------------------------------
CC Copyrighted by the UniProt Consortium, see https://www.uniprot.org/terms
CC Distributed under the Creative Commons Attribution (CC BY 4.0) License
CC ---------------------------------------------------------------------------
DR EMBL; CP047616; QIW54544.1; -; Genomic_DNA.
DR RefSeq; WP_167839034.1; NZ_CP047616.1.
DR AlphaFoldDB; A0A6H0UGI6; -.
DR Proteomes; UP000501945; Chromosome.
DR GO; GO:0003677; F:DNA binding; IEA:UniProtKB-KW.
DR GO; GO:0004520; F:DNA endonuclease activity; IEA:InterPro.
DR GO; GO:0016787; F:hydrolase activity; IEA:UniProtKB-KW.
DR GO; GO:0046872; F:metal ion binding; IEA:UniProtKB-UniRule.
DR GO; GO:0051607; P:defense response to virus; IEA:UniProtKB-UniRule.
DR GO; GO:0043571; P:maintenance of CRISPR repeat elements; IEA:UniProtKB-UniRule.
DR Gene3D; 1.20.120.920; CRISPR-associated endonuclease Cas1, C-terminal domain; 1.
DR HAMAP; MF_01470; Cas1; 1.
DR InterPro; IPR050646; Cas1.
DR InterPro; IPR002729; CRISPR-assoc_Cas1.
DR InterPro; IPR042206; CRISPR-assoc_Cas1_C.
DR InterPro; IPR019855; CRISPR-assoc_Cas1_NMENI.
DR NCBIfam; TIGR00287; cas1; 1.
DR NCBIfam; TIGR03639; cas1_NMENI; 1.
DR PANTHER; PTHR34353; CRISPR-ASSOCIATED ENDONUCLEASE CAS1 1; 1.
DR PANTHER; PTHR34353:SF2; CRISPR-ASSOCIATED ENDONUCLEASE CAS1 1; 1.
DR Pfam; PF01867; Cas_Cas1; 1.
PE 3: Inferred from homology;
KW Antiviral defense {ECO:0000256|ARBA:ARBA00023118, ECO:0000256|HAMAP-
KW Rule:MF_01470};
KW DNA-binding {ECO:0000256|ARBA:ARBA00023125, ECO:0000256|HAMAP-
KW Rule:MF_01470};
KW Endonuclease {ECO:0000256|ARBA:ARBA00022759, ECO:0000256|HAMAP-
KW Rule:MF_01470};
KW Hydrolase {ECO:0000256|ARBA:ARBA00022801, ECO:0000256|HAMAP-Rule:MF_01470};
KW Magnesium {ECO:0000256|ARBA:ARBA00022842, ECO:0000256|HAMAP-Rule:MF_01470};
KW Manganese {ECO:0000256|ARBA:ARBA00023211, ECO:0000256|HAMAP-Rule:MF_01470};
KW Metal-binding {ECO:0000256|ARBA:ARBA00022723, ECO:0000256|HAMAP-
KW Rule:MF_01470};
KW Nuclease {ECO:0000256|ARBA:ARBA00022722, ECO:0000256|HAMAP-Rule:MF_01470}.
FT BINDING 150
FT /ligand="Mn(2+)"
FT /ligand_id="ChEBI:CHEBI:29035"
FT /evidence="ECO:0000256|HAMAP-Rule:MF_01470"
FT BINDING 207
FT /ligand="Mn(2+)"
FT /ligand_id="ChEBI:CHEBI:29035"
FT /evidence="ECO:0000256|HAMAP-Rule:MF_01470"
FT BINDING 222
FT /ligand="Mn(2+)"
FT /ligand_id="ChEBI:CHEBI:29035"
FT /evidence="ECO:0000256|HAMAP-Rule:MF_01470"
SQ SEQUENCE 302 AA; 34688 MW; DDE555FF6978A010 CRC64;
Query Match 95.5%; Score 1230; Length 302;
Best Local Similarity 97.2%;
Matches 242; Conservative 0; Mismatches 7; Indels 0; Gaps 0;
Qy 1 MRDIIYVENRYFISSRENALRFHDYINKATHFIAFDDIDILVLDNARSYLSNGVINDCLD 60
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 1 MRDIIYVENRYFISSRENALRFHDYINKATHFIAFDDIDILVLDNARSYLSNGVINDCLD 60
Qy 61 RNILILTCDNKHSPKAILSNAFANKKRLERLRNQLQLSSKSKNRLWRKIVMAKINNQADA 120
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 61 RNILILTCDNKHSPKAILSNAFANKKRLERLRNQLQLSSKSKNRLWRKIVMAKINNQADA 120
Qy 121 VTFTVQDGTVHREIIELGKMVTEGDKDNREAVVARKYFRTLFGGNFKRGRFDDVINSALN 180
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 121 VTFTVQDGTVHREIIELGKMVTEGDKDNREAVVARKYFRTLFGGNFKRGRFDDVINSALN 180
Qy 181 YGYALVRAVIRRELAICGFEMSFGIHHMSTENPFNLSDDMIEVFRPFVDVLVFEIIVTNY 240
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Db 181 YGYALVRAVIRRELAICGFEMSFGIHHMSTENPFNLSDDMIEVFRPFVDVLVFEIIVTND 240
Qy 241 KRKFKRAIK 249
| ||
Db 241 VTVFDYEIK 249
Claim 15 is excluded from this rejection because it is drawn to a method comprising the active steps of introducing into a bacterial strain the polynucleotide of claim 2, which is not a judicial exception.
Claim Rejections - 35 USC § 102
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)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(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 1-11 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Genbank Accession No. CP047616.1 (published 4/9/2020, hereinafter CP047616.1)
Regarding claims 1 and 2, CP047616.1 teaches the genomic sequence of Pseudolactoccus raffinolactis strain Lr_19_5 a polynucleotide, which comprises genes encoding both cas1 and cas2 polypeptides (see attached). It is relevant to note that the cas1 and cas2 sequences from UniProt, shown in the alignments above are amino acid sequences from the same bacterial strain and genome sequence, GenBank Accession No. CP047616.1.
Regarding claim 3, CP047616.1 teaches the polynucleotide encoding a cas2 polypeptide, as described above.
Regarding claims 4 and 6, CP047616.1 teaches that the cas1 polypeptide comes from a Lactococcus bacterial strain, as described above.
Regarding claim 5, CP047616.1 teaches the polynucleotide encoding a cas1 amino acid sequence with 100% identity to SEQ ID NO: 2, as described above.
Regarding claim 7, CP047616.1 teaches the polynucleotide also encoding a cas2 amino acid sequence with 100% identity to SEQ ID NO: 4, as described above.
Regarding claims 8-10, insofar as CP047616.1 and fragments thereof are interpreted as a vector, CP047616.1 teaches vectors comprising the polynucleotide, as described above.
Regarding claim 11, CP047616.1 teaches a bacterial strain, P. raffinolactis (formerly named Lactococcus raffinolactis; see above) comprising the polynucleotide.
Claims 1-3, 8-10 and 11-12 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Sampson (cited above).
Regarding claims 1-3, Sampson teaches bacterial strains comprising polynucleotides (genes) encoding cas1 and cas2 polypeptides (pp. 1619-20):
Analysis of the genome of highly virulent F. tularensis (strain SchuS4) provides strong evidence for the degeneration of its CRISPR/Cas systems compared with F. novicida (strain U112). Specifically, there are disruptions within all four cas genes. While F. tularensis encodes the full-length DNA sequence for cas1, it contains a single base deletion (thymine 556 [815 478]) resulting in a -1 frame-shift mutation, leading to truncation of the protein by 125 amino acids (Fig. 1D). Similarly, this truncation of the cas1 gene is also present in F. holarctica (strain LVS) and F. mediasiatica (strain FSC147). The cas2 gene of F. tularensis contains a single base insertion (adenine 83 [816 119–816 120]) resulting in a +1 frame-shift mutation and a Cas2 protein only 31 amino acids in length, compared with 98 in F. novicida (Fig. 1E), whereas the cas2 open reading frames in F. holarctica and F. mediasiatica appear to be full-length in comparison to F. novicida. Since cas1 is likely nonfunctional in F. tularensis and other virulent Francisella species, these species would lack the ability to integrate new spacer sequences and therefore to adapt to new target sequences.
Regarding claims 8-10, insofar as Sampson teaches the gene or genome comprising the cas1 and cas2 coding sequences, they teach the vectors, as described above.
Regarding claims 11-12, Sampson teaches wherein the bacterial strain is non-adapting, i.e., comprises a degenerated CRISPR/Cas system, as described above.
Claims 1-3, 8-12 and 15 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by He (He et al. Cas1 and Cas2 From the Type II-C CRISPR-Cas System of Riemerella anatipestifer Are Required for Spacer Acquisition. Front. Cell. Infect. Microbiol., 11 June 2018.).
Regarding claims 1-3 and 8-10, He teaches polynucleotides/vectors (plasmids) encoding cas1 and encoding cas1+cas2 (Table 1):
PNG
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133
699
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Greyscale
Regarding claims 11-12 and 15, He teaches a method of enabling adaptation in a non-adapting CRISPR-Cas system, the method comprising introducing into a bacterial strain comprising a non-adapting CRISPR-Cas system (i.e., a Δcas1-cas2 deletion mutant of Riemerella anatipestifer) a polynucleotide of claim 2 (i.e., a polynucleotide encoding a cas1 and a cas2 amino acid sequence, the pET30a plasmid), thus producing the bacterial strain comprising the cas1-cas2 plasmid, as recited in claim 11:
To determine the role of RA-CH-2 Cas1 and Cas2, the mutant strains RA CH-2 Δcas1, RA-CH-2 Δcas2, and RA-CH-2 Δ1cas1-cas2 were constructed…the PCR products amplified from the mutant strains had sizes corresponding to the unexpanded CRISPR1 array (∼200 bp) (Figure 4, Lanes 2–4), indicating the absence of CRISPR expansion in the mutant strains. Spacer acquisition of the cas1 or cas2 deletion mutant was rescued by the introduction of the shuttle plasmid expressing both Cas1 and Cas2 (p. 7)
As with the adaptation of E. coli and S. thermophilus CRISPR-Cas systems or other subtype systems, both Cas1 and Cas2 were required for the adaptation of RA CRISPRCas system, and deletion of either cas1 or cas2 from RACH- 2 abrogated spacer acquisition (p. 9)
Please note that the term “non-adapting CRISPR-Cas system” is interpreted, per the definition provided on p. 14, ln 1-3 of the instant specification, as a “CRISPR-Cas system” which “may not encode or encode non-functioning proteins…that participate in and/or are required for adaptation”. The broadest reasonable interpretation of the term encompasses both naturally occurring and engineered non-adapted systems. Under that interpretation, and because they lack the cas1 and cas2 proteins required for spacer integration, the cas1-cas2 deletion mutants taught by He comprise a non-adapting CRISPR-Cas system.
Claim Rejections - 35 USC § 112(a) – Scope of Enablement
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claim 15 is rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, because the specification, while being enabling for methods of enabling adaptation in a non-adapting CRISPR-Cas system in L. cremoris, comprising introducing a polynucleotide encoding the cas1-cas2 genes from L. raffinolactis, does not reasonably provide enablement for methods of enabling adaptation throughout the scope of the entire genus of any nonadapting CRISPR-Cas system in any recipient strain, comprising introducing a polynucleotide encoding any cas1/cas2 genes from any donor strain. The specification does not enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to use the invention commensurate in scope with these claims.
Factors to be considered in determining whether a disclosure meets the enablement requirement of 35 U.S.C. 112, first paragraph, have been described by the court in In re Wands, 8 USPQ2d 1400 (CA FC 1988). Wands states at page 1404,
“Factors to be considered in determining whether a disclosure would require undue experimentation have been summarized by the board in Ex part Forman. They include (1) the quantity of experimentation necessary, (2) the amount of direction or guidance presented, (3) the presence or absence of working examples, (4) the nature of the invention, (5) the state of the prior art, (6) the relative skill of those in the art, (7) the predictability or unpredictability of the art, and (8) the breadth of the claims.”
The nature of the invention
The invention is the class of invention that the CAFC has characterized as “the unpredictable arts such as chemistry and biology”. Mycogen Plant Sci., Inc. v. Monsanto Co., 243 F. 3d 1316 (Fed. Cir. 2001).
The breadth of the claims
The claimed method encompasses enabling adaptation in any non-adapting CRISPR-Cas system. It comprises introducing a polynucleotide encoding a generic cas1 and generic cas2 polypeptide (of claim 2) into any bacterial strain. The donor and recipients are generic, and therefore may be any bacterium within the Domain of Bacteria. The claim does not state that the CRISPR-Cas system must be endogenous to either organism, so the CRISPR-Cas systems may be any type of system. Since CRISPR-Cas systems are found in prokaryotes from both Archaea and Bacteria, the claim encompasses systems found in either domain. The genus of any prokaryote from Archaea or Bacteria is vast. As evidenced by Jackson, the genus of CRISPR-Cas systems is correspondingly vast, because these systems are widely distributed among prokaryotes, “occurring in 50 and 87% of complete bacterial and archaeal genomes, respectively.” (p. 1)(Jackson et al. CRISPR-Cas: Adapting to change. Science 356, 40 (2017).).
Guidance in the specification and working examples
The specification discloses the discovery of a type III-A CRISPR-Cas system in L. raffinolactis, which was related to that in L. cremoris (p. 76 ln 3-4). Both organisms are bacteria in the genus Lactococcus. Therefore, the ordinary artisan would reasonably have expected that cas enzymes which function in one type III-A CRISPR-Cas system in one bacterial species would function in another, closely related type III-A CRISPR-Cas system in the same or a closely related species (i.e., a bacterium from the same genus). As shown in Table E3 (pp. 76-77), the polypeptides encoded in both CRISPR-Cas loci shared at least 93% identity. The specification also discloses that while complementation of L. raffiolactis Lr_19_5 cas1 or cas2 individually did not result in observable spacer acquisition in L. cremoris DGCC12607 (p. 78 ln 24-29), complementation with both cas1 and cas2 was successful (pp. 77-78). However, E. italicus cas1-cas2 complementation did not yield space acquisition in L. cremoris (p. 79 ln 3-5). The specification discloses that the E. italicus cas1/cas2 share 46.4/63.4% amino acid identity with L. raffinolactis cas1/cas2 (p. 78 ln 33, p. 79 ln 1). Therefore, the specification evidences that while adaptation could be enabled by transferring the cas1 and cas2 genes from one lactococcal strain with a type III-A CRISPR-Cas system into another lactococcal strain with a type III-A CRISPR-Cas system, where the components of the systems had a high percent identity with one another, adaptation was not enabled when transferring a more distantly related, lower-identity set of cas genes.
The unpredictability of the art and the state of the prior art
The state of the art evidences that CRISPR-Cas systems are diverse, variable, and unpredictable, particularly with regard to their cross-functionality (i.e., whether the spacer acquisition components of one system can be transferred from one bacterial genus or species into a different bacterial genus or species which comprises components of a different system, and the resulting hybrid system would still reasonably be expected to have the ability to acquire spacers). As described above, the specification itself indicates that enabling adaptation in one organism using cas1/cas2 enzymes from another, distantly related organism was not successful. The disclosures of the prior art are consistent with this outcome.
In a review of CRISPR adaptation, Jackson stated, “Although many studies have explored CRISPR adaptation in a broad range of host-specific and metagenomic contexts,much of the mechanistic detail has been gleaned from studying a relatively small subset of systems. Thus, despite the relative wealth of mechanistic information about CRISPR adaptation in a few specific types, work in other systems continues to reveal distinct modes of operation for spacer acquisition. Therefore, studies of CRISPR adaptation in alternative systems are necessary to determine which processes are conserved and which are system-specific.” (Review Summary: Outlook). While, “the proteins primarily responsible for catalyzing spacer acquisition – namely, Cas1 and Cas2 – remain relatively conserved, and the genes encoding these proteins are associated with nearly all CRISPR-Cas systems” (p. 1), but, “despite being near ubiquitous among CRISPR-Cas types, Cas1-Cas2 homologs meet the varied requirements for the acquisition of appropriate spacer sequences in different systems. For example, the effector complexes of several CRISPR-Cas types only recognize targets containing a specific sequence adjacent to where the CRISPR RNA (crRNA) base-pairs with the target strand of a MGE (Fig. 1) (31). The crRNA-paired target sequence is termed the protospacer, and the adjacent target-recognition motif is called a protospacer-adjacent motif (PAM)…yet canonical PAM sequences vary between and sometimes within systems.” (Id.), and, “in some systems, such as type III, the length of spacers found within CRISPR arrays appears more variable, and studies of Cas1-Cas2 structure and function in these systems are lacking.”(p. 2). This indicates that Cas1-Cas2 complexes would not have predictably been transferable from one distantly related system to another, given the system-to-system variability in PAM requirements and spacer length.
In some CRISPR-Cas systems, the Cas1-Cas2 complex alone either is known not to be sufficient for spacer integration, or it is unclear what other components are required, and to what extent. Jackson states that, “Although Cas1 and Cas2 play a central role in CRISPR adaptation, type-specific variations in cas gene clusters occur. In many systems, Cas1-Cas2 is assisted by accessory Cas proteins, which are often mutually exclusive and type-specific” (p. 6). This type specificity and mutual exclusivity would have led the ordinary artisan to doubt that the expression of donor Cas1-Cas2 in a recipient would, by itself, have enabled adaptation in situations where the Cas1-Cas2 complex is not native to the recipient system and it requires accessory proteins not present in the recipient.
This lack of transferability exists even in closely related systems, according to Jackson: “Other host proteins may also be necessary for prespacer substrate production. For example, RecG is required for efficient primed CRISPR adaptation in type I-E and I-F systems, but its precise role remains speculative (38, 90). Additionally, it is still enigmatic why some CRISPR-Cas systems require accessory proteins, whereas closely related types do not. For example, type II-C systems lack cas4 and csn2, which assist CRISPR adaptation in type II-A and II-B systems, respectively. These type specific differences exemplify the diversity that has arisen during the evolution of CRISPR-Cas systems.”.(p. 6).
Taken altogether, the instant specification and the prior art indicate that there is a high level of diversity, variability and unpredictability among CRISPR-Cas systems and the components required for spacer acquisition. Given those factors, it would have been difficult to predict what elements are strictly required for spacer integration in a given system and a given prokaryote species, much less whether those elements would successfully enable adaptation in other, less closely related systems and species.
The quantity of experimentation
The quantity of experimentation would be high. It would require orthogonally testing Cas1-Cas2 expression vectors throughout the genus of CRISPR-Cas systems, in combination, to determine which Cas1-Cas2 complexes are cross-compatible with which CRISPR-Cas systems and which are not. It would also require first discovering what other components are required for spacer integration in all of those systems. As described above, given the diversity and variability of CRISPR-Cas systems and the lack of clarity about how space integration functions in all of those systems, the additional experimentation would require de novo experimentation without a guarantee of success. Further considering that any positive results (i.e., discovering what additional enzymes are required for spacer integration in systems where that is unknown) would amount to a significant advancement in the state of the art, the additional experimentation required is considered undue.
Conclusion
Taking into consideration the factors outlined above, including the nature of the invention, the breadth of the claims, the state of the art, the guidance provided by the applicant and the specific examples, it is the conclusion that an undue amount experimentation would be required to make and use the invention as claimed.
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 (i.e., changing from AIA to pre-AIA ) 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 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 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 nonobviousness.
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.
Claim 13 is rejected under 35 U.S.C. 103 as being unpatentable over He (cited above) in view of Kolesnik (Kolesnik et al. Type III CRISPR-Cas Systems: Deciphering the Most Complex Prokaryotic Immune System. Biochemistry (Mosc). 2021 Oct 20;86(10):1301–1314.)
He teaches the bacterial strain with a non-adapting CRISPR-Cas system, comprising a polynucleotide of claim 2 (comprising cas1 and cas2 coding sequences), as recited in claim 11 from which claim 13 depends.
He does not teach that the CRISPR-Cas system is a type III-A system.
However, He does provide a motivation to delete endogenous cas1 and cas2 genes in a bacterium, followed by partial or full rescue by administration of a plasmid expressing those genes, to study the role of those proteins in spacer acquisition in a given bacterium (see e.g., Abstract, p. 7).
Kolesnik teaches that type III CRISPR-Cas systems are among the most common CRISPR-Cas systems, but are also some of the least investigated, mostly due to their complexity (Abstract). Kolesnik further teaches that some type III CRISPR-Cas systems employ cas1-cas2 integration complexes for spacer acquisition, while others encode both cas2 and reverse transcriptase domains fused with or adjacent to cas1, and that the mechanisms responsible for spacer acquisition in those systems are not well-understood (p. 1308).
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 to have modified the polynucleotide encoding cas1 and cas2 to study spacer acquisition in a type II-C CRISPR-Cas system, as taught by He, to encode cas1/cas2 polypeptides from type III-A CRISPR-Cas systems to perform similar experiments in type III-A systems, as taught by Kolesnik. The ordinary artisan would have been motivated to perform by the combined teachings of He and Kolesnik that experiments involving deletion and complementation of the genes involved in CRISPR spacer acquisition could be informative regarding the roles of those genes in various CRISPR-Cas systems (He), and that there was an art-recognized need for further study of type III CRISPR-Cas systems (including type III-A)(Kolesnik). He’s approaches used standard molecular biology techniques (gene deletion, gene transfer via plasmid), which would have given the ordinary artisan a reasonable expectation that they could have been applied to other bacterial strains and systems.
Note: In comparing the claims to the prior art, it is noted that while the claims were previously rejected in this Office Action for lack of enablement, the claims are rejected here insofar as the prior art teaches at least some embodiments encompassed by the claims. However, the application as filed is not particularly enabling for, nor does it specifically describe, the methods taught by the prior art where the application does not principally contemplate or describe those elements as provided in the prior art. Additionally, it is noted that the particular embodiments of the prior art are not enabling for the generic breadth of the methods as claimed.
Conclusion
No claims are allowed at this time.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to AMANDA M ZAHORIK whose telephone number is (703)756-1433. The examiner can normally be reached M-F 8:00-16:00 EST.
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/AMANDA M ZAHORIK/Examiner, Art Unit 1636
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