LOCUS       sequence-170455-        4346 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of anti-CRISPR protein from Type I-E.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4346)
  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 4346)
  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: pEJS427-pCSDest2-AcrE2
FEATURES             Location/Qualifiers
     source          1..4346
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      719..1174
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(806..825)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1016..1037
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1200..1304
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1305..1373,1374..2165)
                     /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     complement(1523..1542)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2336..2924
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2825..2844
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3078..3095
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3212..3233
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3248..3253,3254..3271,3272..3278)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3286..3302
                     /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)."
     primer_bind     3291..3313
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3310..3326
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3310..3326
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3345..3365
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3347..3365
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     polyA_signal    3478..3612
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3515..3534)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3569..3588
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        complement(3957..3975)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3958..3975)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     promoter        complement(join(4010..4346,1..648))
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 gtcctattga cgtcaatggg cattgagcca ggcgggccat ttaccgtaat tgacgtcaat
       61 gggggaggcg ccatatacgt caataggacc gcccatatga cgtcaatagg aaagaccata
      121 tatagagacc attgacgtca atgggggagt ggctatgggc ggtattagga agccccatat
      181 atggtatatg ggaccgccca ttgggagggg ctatctacgt caataggaaa acccatatat
      241 ggaatactat atggcatagg gccaatacat agtattgaac ctggccaata gccatattgg
      301 catagggcca tattggatat tgcctatata ttgatcctgg catatagcca atatggccgc
      361 cattattggc accatgccaa ttcaatatgg cggacttggc acagggccaa gacccctcct
      421 caaacttggc accgtgccaa gtagacccct ccccagttgg cacagtgcca ggtccgccat
      481 attgaattgg catggtgcca agtccgccat attgaattgg ctggggtcca gatccaccat
      541 attgaattgg cacgagtcca tatacgccat attgaattgg cacaggtcca gatccaccat
      601 attgaattgg catgagtcca tatacgccat attgaattgg cggtcgactg gcgtaatagc
      661 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatgggac
      721 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
      781 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
      841 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
      901 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
      961 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     1021 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     1081 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     1141 gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc
     1201 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     1261 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     1321 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     1381 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     1441 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     1501 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     1561 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     1621 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     1681 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     1741 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     1801 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     1861 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     1921 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     1981 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     2041 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     2101 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     2161 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     2221 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     2281 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     2341 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     2401 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     2461 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     2521 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     2581 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     2641 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     2701 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     2761 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     2821 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     2881 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     2941 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     3001 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     3061 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca
     3121 aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg
     3181 actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac
     3241 cccaggcttt acactttatg ctcccggctc gtatgttgtg tggaattgtg agcggataac
     3301 aatttcacac aggaaacagc tatgaccatg attacgccaa gcgcgcaatt aaccctcact
     3361 aaagggaaca aaagctgggt accgggccca atgcattggc gccgcggccg cgaattaaaa
     3421 aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac
     3481 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     3541 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     3601 catgtctgga tctacgtaat acgactcact atagttctag aggctcgagc tagagttcca
     3661 ttacatgaaa cggatagcca acggtcgtct ccgtgcggtt agtgccgtgc aggtggtggt
     3721 aggcctggaa catgccactt ccgttactct ccccggtttc acggtatacg aggccctcga
     3781 attcctcgcc aagaatctgt tgggctgcgc tcttcgcggc ggctgtaatg tcgacagcgt
     3841 caaccgtgaa gcgctcgccg gagttgtcgt tgtttttgcg ggggtcgatg agataggtat
     3901 tcatggtggc ggcttcgaat cgatgggatc ctgcaaaaag aacaagtagc ttgtattcta
     3961 tagtgtcacc taaatcaagc ttgctccgac gtccccaggc agaatggcgg ttccctaaac
     4021 gagcattgct tatatagacc tcccattagg cacgcctacc gcccatttac gtcaatggaa
     4081 cgcccatttg cgtcattgcc cctccccatt gacgtcaatg gggatgtact tggcagccat
     4141 cgcgggccat ttaccgccat tgacgtcaat gggagtactg ccaatgtacc ctggcgtact
     4201 tccaatagta atgtacttgc caagttacta ttaatagata ttgatgtact gccaagtggg
     4261 ccatttaccg tcattgacgt caataggggg cgtgagaacg gatatgaatg ggcaatgagc
     4321 catcccattg acgtcaatgg tgggtg
//
