LOCUS       sequence-354868-        5747 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector for Cre- and Flp-dependent transgene expression (CRE-ON /
            FLP-OFF) 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 5747)
  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 5747)
  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-VTKD5-GFP
FEATURES             Location/Qualifiers
     source          1..5747
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    51..275
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   277..417
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   277..406
                     /label=AAV2 ITR
     rep_origin      492..947
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(579..598)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     789..810
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(964..983)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1083..1105
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1143..1161)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1229..1333
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1334..1402,1403..2194)
                     /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(1552..1571)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2365..2953
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2854..2873
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   3015..3144
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     intron          3887..4019
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     4002..4021
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     protein_bind    4062..4095
                     /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(4102..4135)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(4186..4219)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(4273..4986,4987..4989,4990..4992))
                     /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(4276..4986,4987..4989,4990..4992))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4318..4339)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     4667..4686
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4926..4947
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      4989..4998
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    5042..5075
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    5126..5159
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    5160..5193
                     /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)"
     misc_feature    join(5202..5747,1..43)
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5255..5275)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5673..5684)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
ORIGIN
        1 cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcctggatc ctgtgccttc
       61 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
      121 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
      181 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
      241 tagcaggcat gctggggatg cggtgggctc tatggcagga acccctagtg atggagttgg
      301 ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac
      361 gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc
      421 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa
      481 gcaaccatag tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
      541 cagcgtgacc gctacacttg ccagcgcctt agcgcccgct cctttcgctt tcttcccttc
      601 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
      661 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc
      721 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
      781 ctttaatagt ggactcttgt tccaaactgg aacaacactc aactctatct cgggctattc
      841 ttttgattta taagggattt tgccgatttc ggtctattgg ttaaaaaatg agctgattta
      901 acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaattttat ggtgcactct
      961 cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc
     1021 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     1081 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa
     1141 gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac
     1201 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     1261 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     1321 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     1381 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     1441 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     1501 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     1561 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     1621 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     1681 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     1741 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     1801 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     1861 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     1921 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     1981 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     2041 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     2101 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     2161 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     2221 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     2281 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     2341 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     2401 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     2461 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     2521 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     2581 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     2641 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     2701 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     2761 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     2821 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     2881 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     2941 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     3001 ttttgctcac atgtcctgca ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc
     3061 gtcgggcgac ctttggtcgc ccggcctcag tgagcgagcg agcgcgcaga gagggagtgg
     3121 ccaactccat cactaggggt tcctgctaga ctgcagaggg cccgcggccg cacgcgttta
     3181 ttcagaatgt tatgcactca cagtggtttg gcatgcatct ggtgaatttt ttttaacgaa
     3241 aaattagtgt tggtttcgat gtatggtagc attctcccta acgtaatttg aataattcag
     3301 caaagcccca ctaccagctg tacttctgca gcctcttcca ttcttttcag cattataatt
     3361 ttggttaatt ttcaatttta ggtcctacgt ctctgcaatt tgtgtatgaa taacagaata
     3421 atttccctct tttgtttcgc ctttcctgtt cctgaatcta aataaagatg gctttttagt
     3481 attaaaagtg gaagaaaatt acaggtaatt atctttgacg gtaaaaacgc tgtaatcagc
     3541 gggctacatg aaaaattact ctaattatgg ctgcatttaa gagaatggaa aaaaaccttc
     3601 ttgtggataa aaaccttaaa ttgtccccaa tgtctgcttc aaattggatg gcactgcagc
     3661 tggaggcttt gttcagaatt gatcctgggg agctacgaac ccaaagtttc acagtaggaa
     3721 gctagatatc gtttaaactt cctgcagccc gggctgggca taaaagtcag ggcagagcca
     3781 tctattgctt acatttgctt ctagcctgca agtcgaggag cgcagccttc cagaagcaga
     3841 gcgcggcgcc ttaagctgca gaagttggtc gtgaggcact gggcacgtaa gtatcaaggt
     3901 tacaagacag gtttaaggag accaatagaa actgggcttg tcgagacaga gaagactctt
     3961 gcgtttctga taggcaccta ttggtcttac tgacatccac tttgcctttc tctccacagg
     4021 tgtccactcc cagttcaatt acagctctta agaaagctag cgaagttcct attctctaga
     4081 aagtatagga acttctccgg aataacttcg tataggatac tttatacgaa gttatgcaga
     4141 atggtagctg gattgtagct gctattagca atatgaaacc tcttaataac ttcgtatagc
     4201 atacattata cgaagttata acgcgccact agtgaattcg agctcggtac ccggggatcc
     4261 tctagagtcg acttacttgt acagctcgtc catgccgaga gtgatcccgg cggcggtcac
     4321 gaactccagc aggaccatgt gatcgcgctt ctcgttgggg tctttgctca gggcggactg
     4381 ggtgctcagg tagtggttgt cgggcagcag cacggggccg tcgccgatgg gggtgttctg
     4441 ctggtagtgg tcggcgagct gcacgctgcc gtcctcgatg ttgtggcgga tcttgaagtt
     4501 caccttgatg ccgttcttct gcttgtcggc catgatatag acgttgtggc tgttgtagtt
     4561 gtactccagc ttgtgcccca ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc
     4621 gatgcggttc accagggtgt cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc
     4681 gtcgtccttg aagaagatgg tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa
     4741 gaagtcgtgc tgcttcatgt ggtcggggta gcggctgaag cactgcacgc cgtaggtcag
     4801 ggtggtcacg agggtgggcc agggcacggg cagcttgccg gtggtgcaga tgaacttcag
     4861 ggtcagcttg ccgtaggtgg catcgccctc gccctcgccg gacacgctga acttgtggcc
     4921 gtttacgtcg ccgtccagct cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc
     4981 cttgctcacc atggtggcgt cgacctgcag gcatgcaagc tttactagtg gtggcgctag
     5041 cataacttcg tataaagtat cctatacgaa gttatttgcc ttaacccaga aattatcact
     5101 gttattcttt agaatggtgc aaagaataac ttcgtataat gtatgctata cgaagttatg
     5161 aagttcctat tctctagaaa gtataggaac ttcggcgcgc caatcaacct ctggattaca
     5221 aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat
     5281 acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct
     5341 ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac
     5401 gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca
     5461 cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca
     5521 tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg
     5581 tggtgttgtc ggggaagctg acgtcctttc catggctgct cgcctgtgtt gccacctgga
     5641 ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt
     5701 cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgcctt
//
