LOCUS       sequence-432137-        5159 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Provides AND intersectional (Cre+Flp) expression of eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5159)
  AUTHORS   Hughes AC, Pollard BG, Xu B, Gammons JW, Chapman P, Bikoff JB, 
            Schwarz LA
  TITLE     A Novel Single Vector Intersectional AAV Strategy for Interrogating 
            Cellular Diversity and Brain Function.
  JOURNAL   bioRxiv. 2023 Feb 8:2023.02.07.527312. doi: 
            10.1101/2023.02.07.527312. Preprint.
  PUBMED    36798174
REFERENCE   2  (bases 1 to 5159)
  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: "bioRxiv. 
            2023 Feb 8:2023.02.07.527312. doi: 10.1101/2023.02.07.527312. 
            Preprint."
COMMENT     Sequence Label: pAAV-hSyn-ConVERGD-eGFP-W3SL
FEATURES             Location/Qualifiers
     source          1..5159
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   3..132
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        166..613
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    722..769
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     protein_bind    complement(782..815)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(908..941)
                     /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(956..1669,1670..1672,1673..1675))
                     /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(959..1669,1670..1672,1673..1675))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1001..1022)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1350..1369
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1609..1630
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1672..1681
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(1796..1843)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     complement(1947..1967)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   2340..2480
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2340..2469
                     /label=AAV2 ITR
     primer_bind     complement(2520..2536)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2520..2536)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2533..2555)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2544..2560
                     /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(2568..2574,2575..2592,2593..2598))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2613..2634
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(2922..3510)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3002..3021)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3681..4472,4473..4541))
                     /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     4304..4323
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4542..4646)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4714..4732
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4770..4792)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4892..4911
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5105..5127
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5119..5136
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5120..5136
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 atcctgcagg cagctgcgcg ctcgctcgct cactgaggcc gcccgggcgt cgggcgacct
       61 ttggtcgccc ggcctcagtg agcgagcgag cgcgcagaga gggagtggcc aactccatca
      121 ctaggggttc ctgcggccgc agatgctgca cgggcccgcg atcgcagtgc aagtgggttt
      181 taggaccagg atgaggcggg gtgggggtgc ctacctgacg accgaccccg acccactgga
      241 caagcaccca acccccattc cccaaattgc gcatccccta tcagagaggg ggaggggaaa
      301 caggatgcgg cgaggcgcgt gcgcactgcc agcttcagca ccgcggacag tgccttcgcc
      361 cccgcctggc ggcgcgcgcc accgccgcct cagcactgaa ggcgcgctga cgtcactcgc
      421 cggtcccccg caaactcccc ttcccggcca ccttggtcgc gtccgcgccg ccgccggccc
      481 agccggaccg caccacgcga ggcgcgagat aggggggcac gggcgcgacc atctgcgctg
      541 cggcgccggc gactcagcgc tgcctcagtc tgcggtgggc agcggaggag tcgtgtcgtg
      601 cctgagagcg cagggtaccg ctagcgaagt tcctattctt caaaaggtat aggaacttca
      661 tcaaaatagg aagaccaatg cttcaccatc gacccgaatt gccaagcatc accatcgaca
      721 cgaagttcct attccgaagt tcctattctc tagaaagtat aggaacttcg tcgaccttaa
      781 gataacttcg tatagcatac attatacgaa gttataaaaa caaaaagaaa agcgcgtttc
      841 gtcctatttg ggactcgtca gctggatgta cgcgtttccg cgaatccagg acgcgcaaag
      901 aaagaaaata acttcgtata gcatacatta tacgaagtta tgaattcatg catgcttact
      961 tgtacagctc gtccatgccg agagtgatcc cggcggcggt cacgaactcc agcaggacca
     1021 tgtgatcgcg cttctcgttg gggtctttgc tcagggcgga ctgggtgctc aggtagtggt
     1081 tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag tggtcggcga
     1141 gctgcacgct gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg atgccgttct
     1201 tctgcttgtc ggccatgata tagacgttgt ggctgttgta gttgtactcc agcttgtgcc
     1261 ccaggatgtt gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg
     1321 tgtcgccctc gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga
     1381 tggtgcgctc ctggacgtag ccttcgggca tggcggactt gaagaagtcg tgctgcttca
     1441 tgtggtcggg gtagcggctg aagcactgca cgccgtaggt cagggtggtc acgagggtgg
     1501 gccagggcac gggcagcttg ccggtggtgc agatgaactt cagggtcagc ttgccgtagg
     1561 tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca
     1621 gctcgaccag gatgggcacc accccggtga acagctcctc gcccttgctc accatggtgg
     1681 cactagtacc ggtctcgagg aagttcctat accttttgaa gaataggaac ttcatcaaaa
     1741 taggaagacc aatgcttcac catcgacccg aattgccaag catcaccatc gacacgaagt
     1801 tcctatactt tctagagaat aggaacttcg gaatagtaac ttccgtacga agctttctag
     1861 agcttagccg attcgatatt cgatgcccga gataatcaac ctctggatta caaaatttgt
     1921 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     1981 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     2041 aaatcctggt tagttcttgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
     2101 acaggggctc ggctgttggg cactgacaat tccgtggtgt ttatttgtga aatttgtgat
     2161 gctattgctt tatttgtaac catctagctt tatttgtgaa atttgtgatg ctattgcttt
     2221 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat
     2281 gtttcaggtt cagggggaga tgtgggaggt tttttaaagc ggccggtaac cggagctcca
     2341 ggaaccccta gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc
     2401 cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg
     2461 agcgcgcagc tgcctgcagg attggatccc aatggcgcgc cgagcttggc gtaatcatgg
     2521 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     2581 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     2641 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     2701 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     2761 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     2821 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     2881 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     2941 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     3001 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     3061 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     3121 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     3181 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     3241 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     3301 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     3361 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     3421 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     3481 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     3541 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     3601 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     3661 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     3721 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     3781 gagggcttac catctggccc cagtgctgca atgataccgc gagaaccacg ctcaccggct
     3841 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     3901 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     3961 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     4021 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     4081 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     4141 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     4201 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     4261 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     4321 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     4381 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     4441 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     4501 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     4561 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     4621 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa
     4681 gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag gcccttttgt
     4741 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagactgtc
     4801 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
     4861 gttggcgggt gtcggggctg gcttaactat gcggcatcag agcagattgt actgagagtg
     4921 caccatatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg catcaggcgc
     4981 cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta
     5041 ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg
     5101 ttttcccagt cacgacgttg taaaacgacg gccagtgaat tgacgcgtat tgggatcca
//
