LOCUS       sequence-272353-        6605 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Intersectional viral expression of BFP in cells expressing both Cre 
            AND Flp.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6605)
  AUTHORS   Fenno LE, Ramakrishnan C, Kim YS, Evans KE, Lo M, Vesuna S, Inoue M,
            Cheung KYM, Yuen E, Pichamoorthy N, Hong ASO, Deisseroth K
  TITLE     Comprehensive Dual- and Triple-Feature Intersectional Single-Vector 
            Delivery of Diverse Functional Payloads to Cells of Behaving 
            Mammals.
  JOURNAL   Neuron. 2020 Jun 20. pii: S0896-6273(20)30432-3. doi: 
            10.1016/j.neuron.2020.06.003.
  PUBMED    32574559
REFERENCE   2  (bases 1 to 6605)
  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: "Neuron. 
            2020 Jun 20. pii: S0896-6273(20)30432-3. doi: 
            10.1016/j.neuron.2020.06.003."
COMMENT     Sequence Label: pAAV-Ef1a-Con/Fon-BFP
FEATURES             Location/Qualifiers
     source          1..6605
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1416..1449)
                     /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(1500..1533)
                     /label=loxN
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AGGTATAC) (Shaw et al., 2021). loxN sites are 
                     compatible with each other, but incompatible with loxP or 
                     lox2272 sites."
     protein_bind    1784..1817
                     /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    1868..1901
                     /label=loxN
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AGGTATAC) (Shaw et al., 2021). loxN sites are 
                     compatible with each other, but incompatible with loxP or 
                     lox2272 sites."
     protein_bind    1908..1941
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     protein_bind    complement(2590..2623)
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     misc_feature    2732..3320
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2785..2805)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3203..3214)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3352..3828
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3551..3570)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3868..4008
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3868..3997
                     /label=AAV2 ITR
     rep_origin      4083..4538
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4170..4189)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4380..4401
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4555..4574)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4674..4696
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4734..4752)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4820..4924
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4925..4993,4994..5785)
                     /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(5143..5162)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5956..6544
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6445..6464
                     /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 gcggccgcac gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gatccataac ttcgtatagg atactttata
     1441 cgaagttatg tagctggagc aatttgaaaa gattgttgta ttacctctag ctgcgcttaa
     1501 taacttcgta tagtatacct tatacgaagt tatactagtc ctcctgtact cacgggtgtg
     1561 gttgatgaag gtcttgctgc cgtagaggaa gctagtagcc aggatgtcga aggcgaaggg
     1621 gagagggccg ccctcgacca ccttgattct catggtctgg gtgccctcgt agggcttgcc
     1681 ttcgccctcg gatgtgcact tgaagtgatg gttgtccacg gtgccctcca tgtacagctt
     1741 catgtgcatg ttctccttaa tcagctcgct catggtggct agcataactt cgtataaagt
     1801 atcctatacg aagttatttg ccttaaccca gaaattatca ctgttattct ttagataggt
     1861 gcaaagaata acttcgtata aggtatacta tacgaagtta tggtaccgaa gttcctattc
     1921 ttcaaatagt ataggaactt caattgttat tggtgcaaag aatcacttcc ttaagtcttt
     1981 atactgccca ggaagttcct attcttcaaa aggtatagga acttcggcgc gccttaatta
     2041 agcttgtgcc ccagtttgct agggaggtcg cagtatctgg ccactgccac ctcgtgctgc
     2101 tcgacgtagg tctcgttgtt ggcctccttg attctttcca gtctgtagtc cacatagtag
     2161 acgccaggca tcttgaggtt cttagcgggt ttcttggatc tatatgtggt cttgatgttt
     2221 gcgatcagat ggctcccgcc cacgagcttc agggccatgt cgtttctgcc ttccaggccg
     2281 ccgtcagcgg ggtacagcgt ctcggtgaag gcctcccagc cgagtgtttt cttctgcatc
     2341 acagggccgt tggatgtgaa gttcacccct ctgatcttga cgttgtagat gaggcagccg
     2401 tcctggaggc tggtgtcctg ggtagcggtc agcacgcccc cgtcttcgta tgtggtgact
     2461 ctctcccatg tgaagccctc agggaaggac tgcttgaaga agtcggggat gccctctgtg
     2521 ggacgacatg acttaaccag ttaaccatgt tctctgaaca tgagaagggc tggccactct
     2581 ccagagctcg aagttcctat actatttgaa gaataggaac ttcttgcctt aacccagaaa
     2641 ttatcactgt tattctttag aatggtgcaa agagaagttc ctataccttt tgaagaatag
     2701 gaacttcgaa ttcgatatca agcttatcga taatcaacct ctggattaca aaatttgtga
     2761 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     2821 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     2881 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     2941 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     3001 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     3061 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     3121 ggggaaatca tcgtcctttc cttggctgct cgcctatgtt gccacctgga ttctgcgcgg
     3181 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     3241 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     3301 cctttgggcc gcctccccgc atcgataccg agcgctgctc gagagatcta cgggtggcat
     3361 ccctgtgacc cctccccagt gcctctcctg gccctggaag ttgccactcc agtgcccacc
     3421 agccttgtcc taataaaatt aagttgcatc attttgtctg actaggtgtc cttctataat
     3481 attatggggt ggaggggggt ggtatggagc aaggggcaag ttgggaagac aacctgtagg
     3541 gcctgcgggg tctattggga accaagctgg agtgcagtgg cacaatcttg gctcactgca
     3601 atctccgcct cctgggttca agcgattctc ctgcctcagc ctcccgagtt gttgggattc
     3661 caggcatgca tgaccaggct cagctaattt ttgttttttt ggtagagacg gggtttcacc
     3721 atattggcca ggctggtctc caactcctaa tctcaggtga tctacccacc ttggcctccc
     3781 aaattgctgg gattacaggc gtgaaccact gctcccttcc ctgtccttct gattttgtag
     3841 gtaaccacgt gcggaccgag cggccgcagg aacccctagt gatggagttg gccactccct
     3901 ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct
     3961 ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc
     4021 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata
     4081 gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     4141 cgctacactt gccagcgcct tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     4201 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     4261 tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg
     4321 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     4381 tggactcttg ttccaaactg gaacaacact caactctatc tcgggctatt cttttgattt
     4441 ataagggatt ttgccgattt cggtctattg gttaaaaaat gagctgattt aacaaaaatt
     4501 taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat
     4561 ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc
     4621 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     4681 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt
     4741 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     4801 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     4861 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     4921 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     4981 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     5041 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     5101 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     5161 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     5221 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     5281 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     5341 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     5401 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     5461 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     5521 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     5581 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     5641 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     5701 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     5761 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     5821 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     5881 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     5941 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     6001 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     6061 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     6121 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     6181 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     6241 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     6301 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     6361 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     6421 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     6481 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     6541 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     6601 catgt
//
