LOCUS       sequence-220500-        3639 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV-T7 promoter expression plasmid for human codon optimized Orf2mor
            anti-CRISPR protein with C-terminal NLS .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3639)
  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 3639)
  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-hOrf2mor-NLS(sv40) (BPK5095)
FEATURES             Location/Qualifiers
     source          1..3639
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             217..237
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             246..263
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(286..303)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    292..516
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(587..603)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(587..603)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(600..622)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    611..627
                     /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(635..641,642..659,660..665))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    680..701
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(818..835)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(989..1577)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1069..1088)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1748..2539,2540..2608))
                     /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     2371..2390
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2609..2713)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2792..2811)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2983..3362
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3363..3566
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3517..3537
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3608..3627
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3608..3626
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      join(3634..3639,1..4)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 atgggcaata acccgaaaac cccagaatac acgcgcaagg ctattagggc atacgagaag
       61 aatttggtga gaaagagcgt tacctttgat gtccgcaagg acgatgatat ggaactgctt
      121 aaaatgatcg aacaagacgg tcgcacattt gctcagatag cacgcaccgc ccttttggaa
      181 caccttcaaa aaggatccgg cggtggaggc agtgggccca agaagaagag gaaagtctga
      241 ccggtcatca tcaccatcac cattgagttt aaacccgctg atcagcctcg actgtgcctt
      301 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
      361 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
      421 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
      481 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct
      541 ggggctcgat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
      601 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
      661 gtaaagccta gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
      721 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
      781 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
      841 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
      901 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
      961 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     1021 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     1081 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     1141 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     1201 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     1261 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     1321 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     1381 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     1441 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     1501 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     1561 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     1621 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     1681 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     1741 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     1801 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     1861 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     1921 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     1981 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     2041 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     2101 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     2161 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     2221 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     2281 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     2341 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     2401 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     2461 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     2521 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     2581 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     2641 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     2701 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
     2761 gatctcccga tcccctaggg tcgactctca gtacaatctg ctctgatgcc gcatagttaa
     2821 gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt
     2881 aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc
     2941 gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga ttattgacta
     3001 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
     3061 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
     3121 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
     3181 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
     3241 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
     3301 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
     3361 tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat
     3421 ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg
     3481 actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac
     3541 ggtgggaggt ctatataagc agagctggtt tagtgaaccg tcagatccgc tagagatccg
     3601 cggccgctaa tacgactcac tatagggaga gccgccacc
//
