LOCUS       sequence-272674-        6607 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Intersectional viral expression of BFP in cells expressing Flp AND 
            NOT Cre.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6607)
  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 6607)
  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-Coff/Fon-BFP
FEATURES             Location/Qualifiers
     source          1..6607
                     /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    1786..1819
                     /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    1870..1903
                     /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    1910..1943
                     /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(2592..2625)
                     /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    2734..3322
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2787..2807)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3205..3216)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3354..3830
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3553..3572)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3870..4010
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3870..3999
                     /label=AAV2 ITR
     rep_origin      4085..4540
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4172..4191)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4382..4403
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4557..4576)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4676..4698
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4736..4754)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4822..4926
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4927..4995,4996..5787)
                     /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(5145..5164)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5958..6546
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6447..6466
                     /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 tatactagtg ccaccatgag cgagctgatt
     1561 aaggagaaca tgcacatgaa gctgtacatg gagggcaccg tggacaacca tcacttcaag
     1621 tgcacatccg agggcgaagg caagccctac gagggcaccc agaccatgag aatcaaggtg
     1681 gtcgagggcg gccctctccc cttcgccttc gacatcctgg ctactagctt cctctacggc
     1741 agcaagacct tcatcaacca cacccgtgag tacaggaggg ctagcataac ttcgtataaa
     1801 gtatcctata cgaagttatt tgccttaacc cagaaattat cactgttatt ctttagatag
     1861 gtgcaaagaa taacttcgta taaggtatac tatacgaagt tatggtaccg aagttcctat
     1921 tcttcaaata gtataggaac ttcaattgtt attggtgcaa agaatcactt ccttaagtct
     1981 ttatactgcc caggaagttc ctattcttca aaaggtatag gaacttcggc gcgccttaat
     2041 taagcttgtg ccccagtttg ctagggaggt cgcagtatct ggccactgcc acctcgtgct
     2101 gctcgacgta ggtctcgttg ttggcctcct tgattctttc cagtctgtag tccacatagt
     2161 agacgccagg catcttgagg ttcttagcgg gtttcttgga tctatatgtg gtcttgatgt
     2221 ttgcgatcag atggctcccg cccacgagct tcagggccat gtcgtttctg ccttccaggc
     2281 cgccgtcagc ggggtacagc gtctcggtga aggcctccca gccgagtgtt ttcttctgca
     2341 tcacagggcc gttggatgtg aagttcaccc ctctgatctt gacgttgtag atgaggcagc
     2401 cgtcctggag gctggtgtcc tgggtagcgg tcagcacgcc cccgtcttcg tatgtggtga
     2461 ctctctccca tgtgaagccc tcagggaagg actgcttgaa gaagtcgggg atgccctctg
     2521 tgggacgaca tgacttaacc agttaaccat gttctctgaa catgagaagg gctggccact
     2581 ctccagagct cgaagttcct atactatttg aagaatagga acttcttgcc ttaacccaga
     2641 aattatcact gttattcttt agaatggtgc aaagagaagt tcctatacct tttgaagaat
     2701 aggaacttcg aattcgatat caagcttatc gataatcaac ctctggatta caaaatttgt
     2761 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     2821 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     2881 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     2941 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     3001 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     3061 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     3121 tcggggaaat catcgtcctt tccttggctg ctcgcctatg ttgccacctg gattctgcgc
     3181 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     3241 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     3301 tccctttggg ccgcctcccc gcatcgatac cgagcgctgc tcgagagatc tacgggtggc
     3361 atccctgtga cccctcccca gtgcctctcc tggccctgga agttgccact ccagtgccca
     3421 ccagccttgt cctaataaaa ttaagttgca tcattttgtc tgactaggtg tccttctata
     3481 atattatggg gtggaggggg gtggtatgga gcaaggggca agttgggaag acaacctgta
     3541 gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt ggcacaatct tggctcactg
     3601 caatctccgc ctcctgggtt caagcgattc tcctgcctca gcctcccgag ttgttgggat
     3661 tccaggcatg catgaccagg ctcagctaat ttttgttttt ttggtagaga cggggtttca
     3721 ccatattggc caggctggtc tccaactcct aatctcaggt gatctaccca ccttggcctc
     3781 ccaaattgct gggattacag gcgtgaacca ctgctccctt ccctgtcctt ctgattttgt
     3841 aggtaaccac gtgcggaccg agcggccgca ggaaccccta gtgatggagt tggccactcc
     3901 ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg
     3961 ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat
     4021 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca
     4081 tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     4141 accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc
     4201 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     4261 tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt
     4321 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     4381 agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat
     4441 ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa
     4501 tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca
     4561 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     4621 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     4681 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
     4741 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     4801 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     4861 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     4921 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     4981 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     5041 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     5101 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     5161 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     5221 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     5281 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     5341 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     5401 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     5461 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     5521 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     5581 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     5641 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     5701 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     5761 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     5821 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     5881 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     5941 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     6001 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     6061 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     6121 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     6181 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     6241 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     6301 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     6361 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     6421 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     6481 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     6541 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     6601 cacatgt
//
