LOCUS       sequence-226953-        4969 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector for CASANOVA (AcrIIA4-LOV2 hybrid) expression in 
            mammalian cells; enables optogenetic control of SpyCas9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4969)
  AUTHORS   Bubeck F, Hoffmann MD, Harteveld Z, Aschenbrenner S, Bietz A, 
            Waldhauer MC, Borner K, Fakhiri J, Schmelas C, Dietz L, Grimm D, 
            Correia BE, Eils R, Niopek D
  TITLE     Engineered anti-CRISPR proteins for optogenetic control of 
            CRISPR-Cas9.
  JOURNAL   Nat Methods. 2018 Oct 30. pii: 10.1038/s41592-018-0178-9. doi: 
            10.1038/s41592-018-0178-9.
  PUBMED    30377362
REFERENCE   2  (bases 1 to 4969)
  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: "Nat 
            Methods. 2018 Oct 30. pii: 10.1038/s41592-018-0178-9. doi: 
            10.1038/s41592-018-0178-9."
COMMENT     Sequence Label: pAAV-CMV-CASANOVA
FEATURES             Location/Qualifiers
     source          1..4969
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(28..456)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(138..159)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     350..369
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     583..605
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     597..614
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     598..614
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        624..642
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   697..826
                     /label=AAV2 ITR
     enhancer        846..1225
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1226..1429
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1380..1400
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        1474..1492
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1518..1538
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(2341..2358)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2347..2571
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2781..2801)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2781..2799)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2820..2836)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2820..2836)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2833..2855)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2844..2860
                     /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(2868..2874,2875..2892,2893..2898))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2913..2934
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3222..3810)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3302..3321)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3981..4772,4773..4841))
                     /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     4604..4623
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4842..4946)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
       61 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
      121 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
      181 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
      241 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
      301 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
      361 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
      421 cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tgcgccattc aggctgcgca
      481 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
      541 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
      601 aaacgacggc cagtgagcgc gcgtaatacg actcactata gggcgaattg gagctctcta
      661 gaatgcaggg gggggggggg ggggsccccy ccctctctgc gcgctcgctc gctcactgag
      721 gccgcccggg caaagcccgg gcgtcgggcg acctttggtc gcccggcctc agtgagcgag
      781 cgagcgcgca gagagggagt ggccaactcc atcactaggg gttcctcaga tctgatatcg
      841 tcgacgacat tgattattga ctagttatta atagtaatca attacggggt cattagttca
      901 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
      961 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
     1021 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
     1081 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
     1141 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
     1201 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
     1261 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
     1321 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
     1381 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac
     1441 tagagaaccc actgcttact ggcttatcga aattaatacg actcactata gggagaccca
     1501 agctggctag catggccccg aagaaaaagc gaaaggtggg cggaagcgga ggttccatga
     1561 acattaacga cctgattcgg gaaataaaga acaaggacta cacggtcaag ctgtcaggca
     1621 ccgactccaa ctctataaca cagctcatta taagggttaa taatgatggc aacgaatatg
     1681 tcatctctga atccgagaat gaatcaatcg tggagaaatt catcagtgca tttaaaaacg
     1741 gctggcttgc tacaactttg gagcggattg agaagaattt cgtcataact gatcccaggt
     1801 tgcctgataa tcctataata ttcgcctccg atagttttct tcaacttacg gagtactcca
     1861 gagaggagat actgggaagg aattgcagat tcctccaagg tccagaaact gatagagcta
     1921 ctgtgaggaa gatacgggat gccattgaca atcaaactga agtgacagtg caacttatca
     1981 attacaccaa gtcagggaaa aaattctgga acctgttcca tttgcaaccc atgagggacc
     2041 aaaagggcga tgtgcaatat ttcataggcg ttcaacttga tggtactgaa cacgtcagag
     2101 atgcggcgga gcgagaagga gtgatgctga ttaagaaaac agcagagaat atcgacgaag
     2161 cggcgaaaga gttgtacgaa gatgaggaag agttctacaa tgatatgcaa actatcacac
     2221 tgaaatctga gctgaactag ctcgagcggc cgccactgtg ctggatatct gcagaattcc
     2281 accacactgg actagtggat ccgagctcgg taccaagctt aagtttaaac cgctgatcag
     2341 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     2401 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     2461 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     2521 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gggcgcgccg
     2581 atatcagatc tgggaaacca gatgatggag gtacccactc cctctatgcg cgctcgctca
     2641 ctcactcggc cctggagacc aaaggtctcc agggcggaga ccaaaggtct ccagggccga
     2701 gtgagtgagc gagcgcgcat agagggagtg ggtaggacgc gtcctgcagg atgcatacta
     2761 gtggtaccca gcttttgttc cctttagtga gggttaattg cgcgcttggc gtaatcatgg
     2821 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     2881 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     2941 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     3001 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     3061 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     3121 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     3181 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     3241 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     3301 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     3361 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     3421 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     3481 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     3541 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     3601 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     3661 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     3721 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     3781 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     3841 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     3901 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     3961 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     4021 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     4081 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     4141 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     4201 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     4261 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     4321 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     4381 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     4441 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     4501 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     4561 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     4621 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     4681 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     4741 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     4801 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     4861 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     4921 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccac
//
