LOCUS       sequence-178072-        5870 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mamalian expression of AcrIIA1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5870)
  AUTHORS   Rauch BJ, Silvis MR, Hultquist JF, Waters CS, McGregor MJ, Krogan 
            NJ, Bondy-Denomy J
  TITLE     Inhibition of CRISPR-Cas9 with Bacteriophage Proteins.
  JOURNAL   Cell. 2016 Dec 22. pii: S0092-8674(16)31683-X. doi: 
            10.1016/j.cell.2016.12.009.
  PUBMED    28041849
REFERENCE   2  (bases 1 to 5870)
  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. 2016 
            Dec 22. pii: S0092-8674(16)31683-X. doi: 
            10.1016/j.cell.2016.12.009."
COMMENT     Sequence Label: pJH372
FEATURES             Location/Qualifiers
     source          1..5870
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        17..396
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        397..600
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     551..571
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     645..664
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        645..663
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1243..1260)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1249..1473
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1519..1947
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1606..1625)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1816..1837
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1956..1976)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1961..2290
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2141..2276
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2203..2222
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2357..3151
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2411..2430)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3021..3040
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3325..3446
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3362..3381)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3416..3435
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3495..3511)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3495..3511)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3508..3530)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3519..3535
                     /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(3543..3549,3550..3567,3568..3573))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3588..3609
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3726..3743)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3897..4485)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3977..3996)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4656..5447,5448..5516))
                     /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     5279..5298
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5517..5621)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5696..5715)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg
       61 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
      121 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
      181 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
      241 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
      301 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
      361 cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc
      421 aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc
      481 aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc
      541 gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct
      601 ctctggctaa ctagagaacc cactgcttac tggcttatcg aaattaatac gactcactat
      661 agggagaccc aagctggcta gcgtttaaac ttaagcttgg taccgagctc ggatccgcca
      721 ccatgaccat caagttgctc gacgaatttc tcaagaaaca tgatcttacc cggtaccaac
      781 tgtctaaact gactggcatt tctcaaaata ccttgaaaga tcagaatgaa aagcccttga
      841 ataagtacac tgtatccata ctcagaagtc tcagcttgat ttccggtctg agcgttagcg
      901 atgtactttt tgaacttgaa gacatagaaa agaattctga tgacttggct ggattcaagc
      961 acctcctcga taaatacaaa ctctccttcc cagcacaaga atttgaactc tactgcctta
     1021 ttaaagaatt cgagtccgca aatatcgaag ttcttccatt cacgtttaat agattcgaga
     1081 acgaggaaca tgtgaatatt aagaaggatg tctgcaaggc tcttgaaaat gccatcaccg
     1141 tcctgaagga aaaaaaaaat gagctgcttt gagaattctg cagatatcca gcacagtggc
     1201 ggccgctcga gtctagaggg cccgtttaaa cccgctgatc agcctcgact gtgccttcta
     1261 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     1321 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     1381 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     1441 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
     1501 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     1561 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     1621 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     1681 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     1741 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     1801 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     1861 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     1921 tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg
     1981 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     2041 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     2101 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     2161 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     2221 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     2281 cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg
     2341 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     2401 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     2461 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     2521 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     2581 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     2641 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     2701 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     2761 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     2821 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     2881 ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     2941 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     3001 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     3061 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     3121 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     3181 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     3241 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     3301 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     3361 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     3421 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     3481 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     3541 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     3601 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     3661 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     3721 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     3781 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     3841 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     3901 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     3961 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     4021 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     4081 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     4141 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     4201 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     4261 aggattagca gagcgaggta tgtaggcagt gctacagagt tcttgaagtg gtggcctaac
     4321 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     4381 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     4441 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     4501 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     4561 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     4621 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     4681 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     4741 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     4801 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     4861 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     4921 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     4981 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     5041 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     5101 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     5161 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     5221 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     5281 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     5341 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     5401 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     5461 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     5521 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     5581 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     5641 ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
     5701 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
     5761 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
     5821 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg
//
