LOCUS       sequence-233705-        5168 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector for expression of AcrIIA4 (no miR binding sites, control 
            vector).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5168)
  AUTHORS   Hoffmann MD, Aschenbrenner S, Grosse S, Rapti K, Domenger C, Fakhiri
            J, Mastel M, Borner K, Eils R, Grimm D, Niopek D
  TITLE     Cell-specific CRISPR-Cas9 activation by microRNA-dependent 
            expression of anti-CRISPR proteins.
  JOURNAL   Nucleic Acids Res. 2019 Apr 15. pii: 5458120. doi: 
            10.1093/nar/gkz271.
  PUBMED    30982889
REFERENCE   2  (bases 1 to 5168)
  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: "Nucleic 
            Acids Res. 2019 Apr 15. pii: 5458120. doi: 10.1093/nar/gkz271."
COMMENT     Sequence Label:  AAV CMV-driven AcrIIA4-scaffold (2xBsmBI sites)
FEATURES             Location/Qualifiers
     source          1..5168
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     enhancer        397..776
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        777..980
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     931..951
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        1025..1043
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1065..1088
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(1481..1498)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1487..1711
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(1922..1940)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1950..1967)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1950..1966)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1959..1981)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(2107..2562)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2244..2265)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2456..2475
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2589..2693
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2694..2762,2763..3554)
                     /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     complement(2912..2931)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3725..4313
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4214..4233
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    4601..4622
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4637..4642,4643..4660,4661..4667)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4675..4691
                     /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)."
     primer_bind     4680..4702
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4699..4715
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4699..4715
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4734..4754
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4736..4754
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg agtcttcgag gaagacaggt ttaagagcta tgctggaaac agcatagcaa
      301 gtttaaataa ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt
      361 tgcggccgcg gccgctctac aaatgtggta tctagagaca ttgattattg actagttatt
      421 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
      481 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
      541 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
      601 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
      661 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
      721 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
      781 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
      841 gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc
      901 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
      961 aggtctatat aagcagagct ctctggctaa ctagagaacc cactgcttac tggcttatcg
     1021 aaattaatac gactcactat agggagaccc aagctggcta gatggactat aaggacgatg
     1081 acgataaagc tggcggcagc atgaacatta acgacctgat tcgggaaata aagaacaagg
     1141 actacacggt caagctgtca ggcaccgact ccaactctat aacacagctc attataaggg
     1201 ttaataatga tggcaacgaa tatgtcatct ctgaatccga gaatgaatca atcgtggaga
     1261 aattcatcag tgcatttaaa aacggctgga accaggagta cgaagatgag gaagagttct
     1321 acaatgatat gcaaactatc acactgaaat ctgagctgaa ctagctcgag cggccgccac
     1381 tgtgctggat atctgcagaa ttagagacgc cgttgatgtt gaagtttata taccgttgat
     1441 gttgaagtcg tctctagctt aagtttaaac cgctgatcag cctcgactgt gccttctagt
     1501 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     1561 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     1621 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     1681 aggcatgctg gggatgcggt gggctctatg ggtcgactcg acgatatcag atctgaggaa
     1741 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccgcc
     1801 cgggcaaagc ccgggcgtcg ggcgaccttt ggtcgcccgg cctcagtgag cgagcgagcg
     1861 cgcagagagg gagtggccnc cccccccccc cccccctgca ttctagagag ctccaattcg
     1921 ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg
     1981 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     2041 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     2101 gaatggaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     2161 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     2221 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     2281 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     2341 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     2401 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     2461 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     2521 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg
     2581 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2641 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     2701 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     2761 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     2821 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     2881 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     2941 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     3001 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     3061 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     3121 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     3181 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     3241 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     3301 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     3361 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     3421 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     3481 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     3541 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     3601 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     3661 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     3721 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     3781 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     3841 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     3901 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     3961 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     4021 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     4081 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     4141 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     4201 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     4261 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4321 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     4381 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     4441 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     4501 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     4561 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     4621 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     4681 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag cgcgcaatta
     4741 accctcacta aagggaacaa aagctgggta ccactagtat gcatcctgca ggacgcgtcc
     4801 tacccactcc ctctatgcgc gctcgctcac tcactcggcc ctggagacca aaggtctcca
     4861 gggccgagtg agtgagcgag cgcgcataga gggagtgggt acctccatca tctggtttcc
     4921 cagatctgat atcggcgcgc ccgagtccaa cacccgtggg aatcccatgg gcaccatggc
     4981 ccctcgctcc aaaaatgctt tcgcgtcgcg cagacactgc tcggtagttt cggggatcag
     5041 cgtttgagta agagcccgcg tctgaaccct ccgcgccgcc ccggccccag tggaaagacg
     5101 cgcaggcaaa acgcaccacg tgacggagcg tgaccgcgcg ccgagcgcgc gccaaggtcg
     5161 ggcaggaa
//
