LOCUS       sequence-170098-        4349 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of Type II-C anti-CRISPR protein AcrIIC1Nme 
            from Neisseria meningitidis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4349)
  AUTHORS   Pawluk A, Amrani N, Zhang Y, Garcia B, Hidalgo-Reyes Y, Lee J, 
            Edraki A, Shah M, Sontheimer EJ, Maxwell KL, Davidson AR
  TITLE     Naturally Occurring Off-Switches for CRISPR-Cas9.
  JOURNAL   Cell. 2016 Dec 15;167(7):1829-1838.e9. doi: 
            10.1016/j.cell.2016.11.017. Epub 2016 Dec 8.
  PUBMED    27984730
REFERENCE   2  (bases 1 to 4349)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cell.2016.11.017"; journalName: "Cell"; date: 
            "2016-12-15- 15"; volume: "167"; issue: "7"; pages: "1829-1838.e9"
COMMENT     Sequence Label: pEJS433-pCSDest2-AcrIIC1Nme
FEATURES             Location/Qualifiers
     source          1..4349
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        860..878
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     860..877
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     polyA_signal    1226..1360
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1250..1269)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1304..1323
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1473..1493)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1473..1491)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1512..1528)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1512..1528)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1525..1547)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1536..1552
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1560..1566,1567..1584,1585..1590))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1605..1626
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1743..1760)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1914..2502)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1994..2013)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2673..3464,3465..3533))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3296..3315
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3534..3638)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3664..4119)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3801..3822)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4013..4032
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(4190..4349,1..825)
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 aattcaatat ggcggacctg gcactgtgcc aactggggag gggtctactt ggcacggtgc
       61 caagtttgag gaggggtctt ggccctgtgc caagtccgcc atattgaatt ggcatggtgc
      121 caataatggc ggccatattg gctatatgcc aggatcaata tataggcaat atccaatatg
      181 gccctatgcc aatatggcta ttggccaggt tcaatactat gtattggccc tatgccatat
      241 agtattccat atatgggttt tcctattgac gtagatagcc cctcccaatg ggcggtccca
      301 tataccatat atggggcttc ctaataccgc ccatagccac tcccccattg acgtcaatgg
      361 tctctatata tggtctttcc tattgacgtc atatgggcgg tcctattgac gtatatggcg
      421 cctcccccat tgacgtcaat tacggtaaat ggcccgcctg gctcaatgcc cattgacgtc
      481 aataggacca cccaccattg acgtcaatgg gatggctcat tgcccattca tatccgttct
      541 cacgccccct attgacgtca atgacggtaa atggcccact tggcagtaca tcaatatcta
      601 ttaatagtaa cttggcaagt acattactat tggaagtacg ccagggtaca ttggcagtac
      661 tcccattgac gtcaatggcg gtaaatggcc cgcgatggct gccaagtaca tccccattga
      721 cgtcaatggg gaggggcaat gacgcaaatg ggcgttccat tgacgtaaat gggcggtagg
      781 cgtgcctaat gggaggtcta tataagcaat gctcgtttag ggaaccgcca ttctgcctgg
      841 ggacgtcgga gcaagcttga tttaggtgac actatagaat acaagctact tgttcttttt
      901 gcaggatccc atcgattcga agccgccacc atgaataaaa cttataaaat tggaaaaaat
      961 gccgggtatg atggctgcgg tctttgtctt gcggccattt ctgaaaatga agctatcaaa
     1021 gttaagtatt tgcgcgacat ttgtcctgat tacgatggcg atgataaagc tgaggattgg
     1081 ctgagatggg gaacggacag ccgcgtcaaa gcagccgctc ttgaaatgga gcagtacgca
     1141 tatacgtcgg ttggtatggc ctcatgttgg gagtttgttg aactatgact cgagcctcta
     1201 gaactatagt gagtcgtatt acgtagatcc agacatgata agatacattg atgagtttgg
     1261 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat
     1321 tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca
     1381 ttttatgttt caggttcagg gggaggtgtg ggaggttttt taattcgcgg ccgcggcgcc
     1441 aatgcattgg gcccggtacc cagcttttgt tccctttagt gagggttaat tgcgcgcttg
     1501 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     1561 aacatacgag ccgggagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     1621 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     1681 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     1741 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     1801 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     1861 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     1921 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     1981 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     2041 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     2101 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     2161 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     2221 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     2281 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     2341 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     2401 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     2461 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     2521 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     2581 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     2641 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     2701 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     2761 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     2821 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     2881 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     2941 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     3001 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     3061 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     3121 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     3181 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     3241 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     3301 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     3361 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     3421 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     3481 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     3541 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     3601 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     3661 cctaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     3721 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     3781 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     3841 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     3901 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     3961 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     4021 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     4081 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca ttcaggctgc
     4141 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag tcgaccgcca
     4201 attcaatatg gcgtatatgg actcatgcca attcaatatg gtggatctgg acctgtgcca
     4261 attcaatatg gcgtatatgg actcgtgcca attcaatatg gtggatctgg accccagcca
     4321 attcaatatg gcggacttgg caccatgcc
//
