LOCUS       sequence-220497-        3951 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV-T7 promoter expression plasmid for human codon optimized 
            AcrVA3.1 anti-CRISPR protein with C-terminal NLS .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3951)
  AUTHORS   Marino ND, Zhang JY, Borges AL, Sousa AA, Leon LM, Rauch BJ, Walton 
            RT, Berry JD, Joung JK, Kleinstiver BP, Bondy-Denomy J
  TITLE     Discovery of widespread Type I and Type V CRISPR-Cas inhibitors.
  JOURNAL   Science. 2018 Sep 6. pii: science.aau5174. doi: 
            10.1126/science.aau5174.
  PUBMED    30190308
REFERENCE   2  (bases 1 to 3951)
  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: "Science. 
            2018 Sep 6. pii: science.aau5174. doi: 10.1126/science.aau5174."
COMMENT     Sequence Label: pCMV-T7-hAcrVA3.1-NLS(sv40) (RTW2624)
FEATURES             Location/Qualifiers
     source          1..3951
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             529..549
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             558..575
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(598..615)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    604..828
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(899..915)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(899..915)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(912..934)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    923..939
                     /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(947..953,954..971,972..977))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    992..1013
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1130..1147)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1301..1889)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1381..1400)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2060..2851,2852..2920))
                     /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     2683..2702
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2921..3025)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3104..3123)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3295..3674
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3675..3878
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3829..3849
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3920..3939
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3920..3938
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      join(3946..3951,1..4)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 atgggcatag cgcatcagaa gaaccggaga gcggattggg aaagcgtaga ctggactaaa
       61 cataatgatg agattgccca actgctgagc agacatcctg atagtgtagc aaagatgaga
      121 acgaagttcg gcgctcaggg aatggctaaa cggaagccga ggcgcaaata taaggtcaca
      181 cgaaaggctg tcccgcctcc tcatacgcag gagctcgcaa cagccgctgc aaaaatctcc
      241 ccaaagagtg gacgctacga aaccaacgtg aacgctaaaa gatggctcat catatccccg
      301 agtggtcaga ggttcgagtt ctccaacctc cagcatttcg tacgcaatca cccggaattg
      361 tttgccaagg cggatacggt ttggaagcgc caggggggaa agaggggtac ggggggcgaa
      421 tattgcaacg cgagcaatgg acttgctcag gccgccaggt tgaacatagg gtggaagggg
      481 tggcaagcga agatcattaa aggcggatcc ggcggtggag gcagtgggcc caagaagaag
      541 aggaaagtct gaccggtcat catcaccatc accattgagt ttaaacccgc tgatcagcct
      601 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
      661 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
      721 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
      781 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg
      841 aaagaaccag ctggggctcg ataccgtcga cctctagcta gagcttggcg taatcatggt
      901 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
      961 gaagcataaa gtgtaaagcc tagggtgcct aatgagtgag ctaactcaca ttaattgcgt
     1021 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     1081 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     1141 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     1201 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     1261 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     1321 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     1381 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     1441 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     1501 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     1561 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     1621 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     1681 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     1741 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     1801 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     1861 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     1921 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     1981 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     2041 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     2101 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     2161 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     2221 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     2281 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     2341 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     2401 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     2461 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     2521 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     2581 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     2641 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     2701 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     2761 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     2821 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     2881 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     2941 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     3001 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg
     3061 gatcgggaga tcgatctccc gatcccctag ggtcgactct cagtacaatc tgctctgatg
     3121 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
     3181 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
     3241 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
     3301 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
     3361 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
     3421 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
     3481 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     3541 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     3601 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
     3661 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
     3721 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
     3781 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
     3841 gtaggcgtgt acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc
     3901 gctagagatc cgcggccgct aatacgactc actataggga gagccgccac c
//
