LOCUS       sequence-170453-        4445 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of Type II-C anti-CRISPR protein AcrIIC3Nme 
            from Neisseria meningitidis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4445)
  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 4445)
  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: pEJS443-pCSDest2-AcrIIC3Nme
FEATURES             Location/Qualifiers
     source          1..4445
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        914..932
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     914..931
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     polyA_signal    1376..1510
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1400..1419)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1454..1473
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1623..1643)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1623..1641)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1662..1678)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1662..1678)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1675..1697)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1686..1702
                     /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(1710..1716,1717..1734,1735..1740))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1755..1776
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1893..1910)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2064..2652)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2144..2163)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2823..3614,3615..3683))
                     /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     3446..3465
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3684..3788)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3814..4269)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3951..3972)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4163..4182
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(4340..4445,1..879)
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 atatggtgga tctggacccc agccaattca atatggcgga cttggcacca tgccaattca
       61 atatggcgga cctggcactg tgccaactgg ggaggggtct acttggcacg gtgccaagtt
      121 tgaggagggg tcttggccct gtgccaagtc cgccatattg aattggcatg gtgccaataa
      181 tggcggccat attggctata tgccaggatc aatatatagg caatatccaa tatggcccta
      241 tgccaatatg gctattggcc aggttcaata ctatgtattg gccctatgcc atatagtatt
      301 ccatatatgg gttttcctat tgacgtagat agcccctccc aatgggcggt cccatatacc
      361 atatatgggg cttcctaata ccgcccatag ccactccccc attgacgtca atggtctcta
      421 tatatggtct ttcctattga cgtcatatgg gcggtcctat tgacgtatat ggcgcctccc
      481 ccattgacgt caattacggt aaatggcccg cctggctcaa tgcccattga cgtcaatagg
      541 accacccacc attgacgtca atgggatggc tcattgccca ttcatatccg ttctcacgcc
      601 ccctattgac gtcaatgacg gtaaatggcc cacttggcag tacatcaata tctattaata
      661 gtaacttggc aagtacatta ctattggaag tacgccaggg tacattggca gtactcccat
      721 tgacgtcaat ggcggtaaat ggcccgcgat ggctgccaag tacatcccca ttgacgtcaa
      781 tggggagggg caatgacgca aatgggcgtt ccattgacgt aaatgggcgg taggcgtgcc
      841 taatgggagg tctatataag caatgctcgt ttagggaacc gccattctgc ctggggacgt
      901 cggagcaagc ttgatttagg tgacactata gaatacaagc tacttgttct ttttgcagga
      961 tcccatcgat tcgaagccgc caccatgttc aaacgcgcta ttatcttcac ttctttcaac
     1021 ggctttgaaa aagtttctcg aactgaaaaa cgccgccttg ccaaaatcat caatgctcga
     1081 gtttccatca tcgacgaata cttgagagcc aaagacacca acgcatcgct tgacggtcag
     1141 taccgcgctt tcttgttcaa cgacgaatcg cccgcaatga ccgaatttct ggcaaaactt
     1201 aaagcctttg ccgaaagttg caccggaatc agcatcgacg catgggaaat tgaagaaagc
     1261 gaatacgtcc gcctgccggt ggaacgcagg gatttcttag cggcagccaa cggcaaagag
     1321 atttttaaaa tttaatgact cgagcctcta gaactatagt gagtcgtatt acgtagatcc
     1381 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     1441 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     1501 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     1561 ggaggttttt taattcgcgg ccgcggcgcc aatgcattgg gcccggtacc cagcttttgt
     1621 tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg
     1681 tgaaattgtt atccgctcac aattccacac aacatacgag ccgggagcat aaagtgtaaa
     1741 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     1801 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     1861 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     1921 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     1981 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     2041 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     2101 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2161 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     2221 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     2281 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     2341 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     2401 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     2461 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     2521 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     2581 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     2641 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     2701 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     2761 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     2821 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     2881 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     2941 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     3001 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     3061 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     3121 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     3181 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     3241 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     3301 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     3361 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     3421 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     3481 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     3541 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     3601 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     3661 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     3721 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     3781 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa
     3841 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     3901 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     3961 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     4021 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     4081 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     4141 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     4201 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     4261 agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     4321 cctcttcgct attacgccag tcgaccgcca attcaatatg gcgtatatgg actcatgcca
     4381 attcaatatg gtggatctgg acctgtgcca attcaatatg gcgtatatgg actcgtgcca
     4441 attca
//
