LOCUS       sequence-346966-        4467 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In vitro transcription of anti-CRISPR AcrIIC3 mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4467)
  AUTHORS   Takasugi PR, Wang S, Truong KT, Drage EP, Kanishka SN, Higbee MA, 
            Bamidele N, Ojelabi O, Sontheimer EJ, Gagnon JA
  TITLE     Orthogonal CRISPR-Cas tools for genome editing, inhibition, and 
            CRISPR recording in zebrafish embryos.
  JOURNAL   Genetics. 2021 Nov 4. pii: 6420709. doi: 10.1093/genetics/iyab196.
  PUBMED    34735006
REFERENCE   2  (bases 1 to 4467)
  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: "Genetics. 
            2021 Nov 4. pii: 6420709. doi: 10.1093/genetics/iyab196."
COMMENT     Sequence Label: pCS2-AcrIIC3
FEATURES             Location/Qualifiers
     source          1..4467
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(26..43)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(197..785)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(277..296)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(956..1747,1748..1816))
                     /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     1579..1598
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1817..1921)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1947..2402)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2084..2105)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2296..2315
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2473..3457
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        3492..3510
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3492..3509
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             3566..3586
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    3976..4110
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4000..4019)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4054..4073
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4223..4243)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4223..4241)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4262..4278)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4262..4278)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4275..4297)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4286..4302
                     /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(4310..4316,4317..4334,4335..4340))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4355..4376
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
       61 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
      121 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
      181 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
      241 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
      301 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
      361 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
      421 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
      481 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
      541 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
      601 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
      661 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
      721 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
      781 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
      841 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
      901 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
      961 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     1021 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     1081 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     1141 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     1201 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     1261 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     1321 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     1381 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     1441 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     1501 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     1561 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     1621 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     1681 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     1741 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     1801 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     1861 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     1921 gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt taaaattcgc
     1981 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     2041 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     2101 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     2161 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     2221 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     2281 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     2341 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     2401 gtcccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     2461 gctattacgc cagtcgaccg ccaattcaat atggcgtata tggactcatg ccaattcaat
     2521 atggtggatc tggacctgtg ccaattcaat atggcgtata tggactcgtg ccaattcaat
     2581 atggtggatc tggaccccag ccaattcaat atggcggact tggcaccatg ccaattcaat
     2641 atggcggacc tggcactgtg ccaactgggg aggggtctac ttggcacggt gccaagtttg
     2701 aggaggggtc ttggccctgt gccaagtccg ccatattgaa ttggcatggt gccaataatg
     2761 gcggccatat tggctatatg ccaggatcaa tatataggca atatccaata tggccctatg
     2821 ccaatatggc tattggccag gttcaatact atgtattggc cctatgccat atagtattcc
     2881 atatatgggt tttcctattg acgtagatag cccctcccaa tgggcggtcc catataccat
     2941 atatggggct tcctaatacc gcccatagcc actcccccat tgacgtcaat ggtctctata
     3001 tatggtcttt cctattgacg tcatatgggc ggtcctattg acgtatatgg cgcctccccc
     3061 attgacgtca attacggtaa atggcccgcc tggctcaatg cccattgacg tcaataggac
     3121 cacccaccat tgacgtcaat gggatggctc attgcccatt catatccgtt ctcacgcccc
     3181 ctattgacgt caatgacggt aaatggccca cttggcagta catcaatatc tattaatagt
     3241 aacttggcaa gtacattact attggaagta cgccagggta cattggcagt actcccattg
     3301 acgtcaatgg cggtaaatgg cccgcgatgg ctgccaagta catccccatt gacgtcaatg
     3361 gggaggggca atgacgcaaa tgggcgttcc attgacgtaa atgggcggta ggcgtgccta
     3421 atgggaggtc tatataagca atgctcgttt agggaaccgc cattctgcct ggggacgtcg
     3481 gagcaagctt gatttaggtg acactataga atacaagcta cttgttcttt ttgcaggatc
     3541 ccatcgattc gaattcgcca ccatgccaaa aaagaagaga aaggtagaag accccatgtt
     3601 caaacgcgct attatcttca cttctttcaa cggctttgaa aaagtttctc gaactgaaaa
     3661 acgccgcctt gccaaaatca tcaatgctcg agtttccatc atcgacgaat acttgagagc
     3721 caaagacacc aacgcatcgc ttgacggtca gtaccgcgct ttcttgttca acgacgaatc
     3781 gcccgcaatg accgaatttc tggcaaaact taaagccttt gccgaaagtt gcaccggaat
     3841 cagcatcgac gcatgggaaa ttgaagaaag cgaatacgtc cgcctgccgg tggaacgcag
     3901 ggatttctta gcggcagcca acggcaaaga gatttttaaa atttaatcta gaactatagt
     3961 gagtcgtatt acgtagatcc agacatgata agatacattg atgagtttgg acaaaccaca
     4021 actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt
     4081 gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt
     4141 caggttcagg gggaggtgtg ggaggttttt taattcgcgg ccgcggcgcc aatgcattgg
     4201 gcccggtacc cagcttttgt tccctttagt gagggttaat tgcgcgcttg gcgtaatcat
     4261 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     4321 ccgggagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     4381 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4441 tcggccaacg cgcggggaga ggcggtt
//
