LOCUS       sequence-267374-        3723 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV and T7 promoter expression plasmid for human codon optimized 
            AcrIIC1 with C-terminal NLS (SV40).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3723)
  AUTHORS   Mahendra C, Christie KA, Osuna BA, Pinilla-Redondo R, Kleinstiver 
            BP, Bondy-Denomy J
  TITLE     Broad-spectrum anti-CRISPR proteins facilitate horizontal gene 
            transfer.
  JOURNAL   Nat Microbiol. 2020 Apr;5(4):620-629. doi: 
            10.1038/s41564-020-0692-2. Epub 2020 Mar 26.
  PUBMED    32218510
REFERENCE   2  (bases 1 to 3723)
  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: 
            "10.1038/s41564-020-0692-2"; journalName: "Nat Microbiol"; date: 
            "2020-04"; volume: "5"; issue: "4"; pages: "620-629"
COMMENT     Sequence Label: pCMV-T7-AcrIIC1-NLS(SV40) (KAC208)
FEATURES             Location/Qualifiers
     source          1..3723
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      652..661
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             958..978
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             987..1004
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1027..1044)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1033..1257
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1328..1344)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1328..1344)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1341..1363)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1352..1368
                     /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(1376..1382,1383..1400,1401..1406))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1421..1442
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1559..1576)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1730..2318)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1810..1829)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2489..3280,3281..3349))
                     /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     3112..3131
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3350..3454)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3533..3552)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc
      601 agatccgcta gagatccgcg gccgctaata cgactcacta tagggagagc cgccaccatg
      661 ggcaaggagg tgttcaagct gaagcccgag ctggtgacct acaagggctg cggctgggcc
      721 ctggcctgca tcaaggacgg cgagatcatc gacctgacct acgtgagaga cctgggcatc
      781 gaggagtacg acgagaactt cgacggcctg gagcccgaga tcatctacta cgacgtggtg
      841 gccagccagg cctgcaagga ggtggcctac agatacgagg agatgggcga gttcaccttc
      901 ggcctgtgca gctgctggga gttcaacgtg atgggatccg gcggtggagg cagtgggccc
      961 aagaagaaga ggaaagtctg accggtcatc atcaccatca ccattgagtt taaacccgct
     1021 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc
     1081 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg
     1141 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca
     1201 agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt
     1261 ctgaggcgga aagaaccagc tggggctcga taccgtcgac ctctagctag agcttggcgt
     1321 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     1381 tacgagccgg aagcataaag tgtaaagcct agggtgccta atgagtgagc taactcacat
     1441 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     1501 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     1561 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     1621 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     1681 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     1741 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     1801 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     1861 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     1921 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     1981 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     2041 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     2101 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     2161 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     2221 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     2281 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     2341 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     2401 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     2461 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     2521 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     2581 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     2641 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     2701 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     2761 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     2821 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     2881 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     2941 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     3001 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     3061 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     3121 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     3181 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     3241 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     3301 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     3361 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     3421 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     3481 acgtcgacgg atcgggagat cgatctcccg atcccctagg gtcgactctc agtacaatct
     3541 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
     3601 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
     3661 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
     3721 gtt
//
