LOCUS       sequence-355033-        5729 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector for Cre- and Flp-dependent transgene expression (CRE-OFF 
            / FLP-ON) of GFP in cortical interneurons under the control of the 
            Dlx enhancer (Dimidschstein et al 2016).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5729)
  AUTHORS   Pouchelon G, Vergara J, McMahon J, Gorissen BL, Lin JD, 
            Vormstein-Schneider D, Niehaus JL, Burbridge TJ, Wester JC, Sherer 
            M, Fernandez-Otero M, Allaway KC, Pelkey K, Chittajallu R, McBain 
            CJ, Fan M, Nasse JS, Wildenberg GA, Fishell G, Dimidschstein J
  TITLE     A versatile viral toolkit for functional discovery in the nervous 
            system.
  JOURNAL   Cell Rep Methods. 2022 May 26;2(6):100225. doi: 
            10.1016/j.crmeth.2022.100225. eCollection 2022 Jun 20.
  PUBMED    35784651
REFERENCE   2  (bases 1 to 5729)
  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 Rep 
            Methods."; date: "2022-05-26"; volume: "2(6):100225. doi"; pages: " 
            10.1016/j.crmeth.2022.100225. eCollection 2022 Jun 20"
COMMENT     Sequence Label: pAAV-VTKD6-GFP
FEATURES             Location/Qualifiers
     source          1..5729
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     intron          873..1005
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     988..1007
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     protein_bind    complement(1048..1081)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    1082..1115
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             complement(join(1250..1963,1964..1966,1967..1969))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(1253..1963,1964..1966,1967..1969))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1295..1316)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1644..1663
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1903..1924
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1966..1975
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(2010..2043)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     protein_bind    complement(2128..2161)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2170..2758
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2223..2243)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2641..2652)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2766..2990
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2992..3132
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2992..3121
                     /label=AAV2 ITR
     rep_origin      3207..3662
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3294..3313)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3504..3525
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3679..3698)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3798..3820
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3858..3876)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3944..4048
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4049..4117,4118..4909)
                     /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(4267..4286)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5080..5668
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5569..5588
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gctagactgc agagggcccg cggccgcacg cgtttattca gaatgttatg
      181 cactcacagt ggtttggcat gcatctggtg aatttttttt aacgaaaaat tagtgttggt
      241 ttcgatgtat ggtagcattc tccctaacgt aatttgaata attcagcaaa gccccactac
      301 cagctgtact tctgcagcct cttccattct tttcagcatt ataattttgg ttaattttca
      361 attttaggtc ctacgtctct gcaatttgtg tatgaataac agaataattt ccctcttttg
      421 tttcgccttt cctgttcctg aatctaaata aagatggctt tttagtatta aaagtggaag
      481 aaaattacag gtaattatct ttgacggtaa aaacgctgta atcagcgggc tacatgaaaa
      541 attactctaa ttatggctgc atttaagaga atggaaaaaa accttcttgt ggataaaaac
      601 cttaaattgt ccccaatgtc tgcttcaaat tggatggcac tgcagctgga ggctttgttc
      661 agaattgatc ctggggagct acgaacccaa agtttcacag taggaagcta gatatcgttt
      721 aaacttcctg cagcccgggc tgggcataaa agtcagggca gagccatcta ttgcttacat
      781 ttgcttctag cctgcaagtc gaggagcgca gccttccaga agcagagcgc ggcgccttaa
      841 gctgcagaag ttggtcgtga ggcactgggc acgtaagtat caaggttaca agacaggttt
      901 aaggagacca atagaaactg ggcttgtcga gacagagaag actcttgcgt ttctgatagg
      961 cacctattgg tcttactgac atccactttg cctttctctc cacaggtgtc cactcccagt
     1021 tcaattacag ctcttaagaa agctagcata acttcgtata gcatacatta tacgaagtta
     1081 tgaagttcct attctctaga aagtatagga acttcgcaga atggtagctg gattgtagct
     1141 gctattagca atatgaaacc tcttagaagt tcctattctt caaaaggtat aggaacttca
     1201 acgcgcccta gaattcgagc tcggtacccg gggatcctct agagtcgact tacttgtaca
     1261 gctcgtccat gccgagagtg atcccggcgg cggtcacgaa ctccagcagg accatgtgat
     1321 cgcgcttctc gttggggtct ttgctcaggg cggactgggt gctcaggtag tggttgtcgg
     1381 gcagcagcac ggggccgtcg ccgatggggg tgttctgctg gtagtggtcg gcgagctgca
     1441 cgctgccgtc ctcgatgttg tggcggatct tgaagttcac cttgatgccg ttcttctgct
     1501 tgtcggccat gatatagacg ttgtggctgt tgtagttgta ctccagcttg tgccccagga
     1561 tgttgccgtc ctccttgaag tcgatgccct tcagctcgat gcggttcacc agggtgtcgc
     1621 cctcgaactt cacctcggcg cgggtcttgt agttgccgtc gtccttgaag aagatggtgc
     1681 gctcctggac gtagccttcg ggcatggcgg acttgaagaa gtcgtgctgc ttcatgtggt
     1741 cggggtagcg gctgaagcac tgcacgccgt aggtcagggt ggtcacgagg gtgggccagg
     1801 gcacgggcag cttgccggtg gtgcagatga acttcagggt cagcttgccg taggtggcat
     1861 cgccctcgcc ctcgccggac acgctgaact tgtggccgtt tacgtcgccg tccagctcga
     1921 ccaggatggg caccaccccg gtgaacagct cctcgccctt gctcaccatg gtggcgtcga
     1981 cctgcaggca tgcaagcttg gtggctagcg aagttcctat actttctaga gaataggaac
     2041 ttcttgcctt aacccagaaa ttatcactgt tattctttag aatggtgcaa agagaagttc
     2101 ctataccttt tgaagaatag gaacttcata acttcgtata gcatacatta tacgaagtta
     2161 tggcgcgcca atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac
     2221 tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt
     2281 gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat
     2341 gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca
     2401 acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc
     2461 cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg
     2521 gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaagctgac gtcctttcca
     2581 tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct
     2641 tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt
     2701 ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcct
     2761 ggatcctgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     2821 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     2881 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     2941 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg caggaacccc
     3001 tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac
     3061 caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca
     3121 gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3181 accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg
     3241 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc ccgctccttt
     3301 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     3361 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     3421 tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     3481 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaactc
     3541 tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct attggttaaa
     3601 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat
     3661 tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca
     3721 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     3781 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     3841 acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     3901 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     3961 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     4021 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     4081 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     4141 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     4201 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     4261 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     4321 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     4381 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     4441 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     4501 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     4561 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     4621 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     4681 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     4741 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     4801 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     4861 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     4921 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     4981 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     5041 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     5101 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     5161 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     5221 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     5281 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     5341 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     5401 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     5461 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     5521 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     5581 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     5641 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     5701 ggccttttgc tggccttttg ctcacatgt
//
