LOCUS       sequence-225082-        4358 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses type II-C anti-CRISPR protein from H. parainfluenzae in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4358)
  AUTHORS   Lee J, Mir A, Edraki A, Garcia B, Amrani N, Lou HE, Gainetdinov I, 
            Pawluk A, Ibraheim R, Gao XD, Liu P, Davidson AR, Maxwell KL, 
            Sontheimer EJ
  TITLE     Potent Cas9 Inhibition in Bacterial and Human Cells by AcrIIC4 and 
            AcrIIC5 Anti-CRISPR Proteins.
  JOURNAL   MBio. 2018 Dec 4;9(6). pii: mBio.02321-18. doi: 
            10.1128/mBio.02321-18.
  PUBMED    30514786
REFERENCE   2  (bases 1 to 4358)
  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"; journalName: "MBio."; 
            date: "2018-12-4"; volume: "9(6). pii: mBio.02321-18. doi"; pages: "
            10.1128/mBio.02321-18"
COMMENT     Sequence Label: pEJS540-pCSDest2-AcrIIC4Hpa
FEATURES             Location/Qualifiers
     source          1..4358
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        56..1040
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1075..1093
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1075..1092
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     polyA_signal    1450..1584
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1474..1493)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1528..1547
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1697..1717)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1697..1715)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1736..1752)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1736..1752)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1749..1771)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1760..1776
                     /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(1784..1790,1791..1808,1809..1814))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1829..1850
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1967..1984)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2138..2726)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2218..2237)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2897..3688,3689..3757))
                     /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     3520..3539
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3758..3862)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3888..4343)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4025..4046)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4237..4256
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagtcga
       61 ccgccaattc aatatggcgt atatggactc atgccaattc aatatggtgg atctggacct
      121 gtgccaattc aatatggcgt atatggactc gtgccaattc aatatggtgg atctggaccc
      181 cagccaattc aatatggcgg acttggcacc atgccaattc aatatggcgg acctggcact
      241 gtgccaactg gggaggggtc tacttggcac ggtgccaagt ttgaggaggg gtcttggccc
      301 tgtgccaagt ccgccatatt gaattggcat ggtgccaata atggcggcca tattggctat
      361 atgccaggat caatatatag gcaatatcca atatggccct atgccaatat ggctattggc
      421 caggttcaat actatgtatt ggccctatgc catatagtat tccatatatg ggttttccta
      481 ttgacgtaga tagcccctcc caatgggcgg tcccatatac catatatggg gcttcctaat
      541 accgcccata gccactcccc cattgacgtc aatggtctct atatatggtc tttcctattg
      601 acgtcatatg ggcggtccta ttgacgtata tggcgcctcc cccattgacg tcaattacgg
      661 taaatggccc gcctggctca atgcccattg acgtcaatag gaccacccac cattgacgtc
      721 aatgggatgg ctcattgccc attcatatcc gttctcacgc cccctattga cgtcaatgac
      781 ggtaaatggc ccacttggca gtacatcaat atctattaat agtaacttgg caagtacatt
      841 actattggaa gtacgccagg gtacattggc agtactccca ttgacgtcaa tggcggtaaa
      901 tggcccgcga tggctgccaa gtacatcccc attgacgtca atggggaggg gcaatgacgc
      961 aaatgggcgt tccattgacg taaatgggcg gtaggcgtgc ctaatgggag gtctatataa
     1021 gcaatgctcg tttagggaac cgccattctg cctggggacg tcggagcaag cttgatttag
     1081 gtgacactat agaatacaag ctacttgttc tttttgcagg atcccatcga ttcgaagccg
     1141 ccaccatgaa gatcaccagc agcaacttcg cgaccattgc gaccagcgag aactttgcga
     1201 agctgagcgt gctgccgaaa aaccaccgtg agccgatcaa gggtctgttc aaaagcgcgg
     1261 ttgaacagtt tagcagcgcg cgtgacttct ttaagaacga gaactacagc aaagagctgg
     1321 cggaaaagtt caacaaagaa gcggtgaacg aggcggttga aaagctgcaa aaagcgatcg
     1381 atctggcgga aaaacagggc attcaatttt gactcgagcc tctagaacta tagtgagtcg
     1441 tattacgtag atccagacat gataagatac attgatgagt ttggacaaac cacaactaga
     1501 atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc
     1561 attataagct gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt
     1621 cagggggagg tgtgggaggt tttttaattc gcggccgcgg cgccaatgca ttgggcccgg
     1681 tacccagctt ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat
     1741 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggga
     1801 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     1861 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     1921 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     1981 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     2041 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     2101 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     2161 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     2221 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     2281 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     2341 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     2401 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     2461 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     2521 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     2581 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     2641 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     2701 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     2761 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     2821 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     2881 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     2941 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     3001 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     3061 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     3121 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     3181 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     3241 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     3301 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     3361 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     3421 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     3481 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     3541 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     3601 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     3661 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     3721 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     3781 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     3841 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt
     3901 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     3961 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
     4021 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     4081 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
     4141 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     4201 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     4261 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     4321 taatgcgccg ctacagggcg cgtcccattc gccattca
//
