LOCUS       sequence-268051-        3897 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV and T7 promoter expression plasmid for human codon optimized 
            AcrIIA1(T114A/F115A) with C-terminal NLS (SV40).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3897)
  AUTHORS   Osuna BA, Karambelkar S, Mahendra C, Christie KA, Garcia B, Davidson
            AR, Kleinstiver BP, Kilcher S, Bondy-Denomy J
  TITLE     Listeria Phages Induce Cas9 Degradation to Protect Lysogenic 
            Genomes.
  JOURNAL   Cell Host Microbe. 2020 Apr 18. pii: S1931-3128(20)30190-6. doi: 
            10.1016/j.chom.2020.04.001.
  PUBMED    32325050
REFERENCE   2  (bases 1 to 3897)
  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: "Cell Host 
            Microbe. 2020 Apr 18. pii: S1931-3128(20)30190-6. doi: 
            10.1016/j.chom.2020.04.001."
COMMENT     Sequence Label: pCMV-T7-AcrIIA1(T114A/F115A)-NLS(SV40) (KAC688)
FEATURES             Location/Qualifiers
     source          1..3897
                     /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             1132..1152
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1161..1178
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1201..1218)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1207..1431
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1502..1518)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1502..1518)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1515..1537)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1526..1542
                     /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(1550..1556,1557..1574,1575..1580))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1595..1616
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1733..1750)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1904..2492)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1984..2003)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2663..3454,3455..3523))
                     /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     3286..3305
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3524..3628)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3707..3726)
                     /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 ggcaccatca agctgctgga cgagttcctg aagaagcacg acctgaccag ataccagctg
      721 agcaagctga ccggcatcag ccagaacacc ctgaaggacc agaacgagaa gcccctgaac
      781 aagtacaccg tgagcatcct gagaagcctg agcctgatca gcggcctgag cgtgagcgac
      841 gtgctgttcg agctggagga catcgagaag aacagcgacg acctggccgg cttcaagcac
      901 ctgctggaca agtacaagct gagcttcccc gcccaggagt tcgagctgta ctgcctgatc
      961 aaggagttcg agagcgccaa catcgaggtg ctgcccttcg ccgccaacag attcgagaac
     1021 gaggagcacg tgaacatcaa gaaggacgtg tgcaaggccc tggagaacgc catcaccgtg
     1081 ctgaaggaga agaagaacga gctgctggga tccggcggtg gaggcagtgg gcccaagaag
     1141 aagaggaaag tctgaccggt catcatcacc atcaccattg agtttaaacc cgctgatcag
     1201 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     1261 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     1321 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     1381 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg
     1441 cggaaagaac cagctggggc tcgataccgt cgacctctag ctagagcttg gcgtaatcat
     1501 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     1561 ccggaagcat aaagtgtaaa gcctagggtg cctaatgagt gagctaactc acattaattg
     1621 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     1681 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     1741 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     1801 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     1861 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     1921 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     1981 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     2041 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     2101 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     2161 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     2221 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     2281 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     2341 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     2401 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     2461 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     2521 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     2581 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     2641 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     2701 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     2761 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     2821 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     2881 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     2941 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     3001 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     3061 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     3121 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     3181 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     3241 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     3301 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     3361 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     3421 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     3481 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     3541 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     3601 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     3661 acggatcggg agatcgatct cccgatcccc tagggtcgac tctcagtaca atctgctctg
     3721 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
     3781 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
     3841 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgtt
//
