LOCUS       sequence-265136-        6624 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Intersectional viral expression of mCherry in cells expressing both 
            Cre AND Flp.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6624)
  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 6624)
  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-mCherry
FEATURES             Location/Qualifiers
     source          1..6624
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        242..1419
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1410
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1367..1387
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1426..1459)
                     /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(1510..1543)
                     /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."
     primer_bind     1742..1760
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      1902..1911
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    1916..1949
                     /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    2000..2033
                     /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    2040..2073
                     /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)"
     primer_bind     complement(2444..2463)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(2609..2642)
                     /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    2751..3339
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2804..2824)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3222..3233)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3371..3847
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3570..3589)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3887..4027
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3887..4016
                     /label=AAV2 ITR
     rep_origin      4102..4557
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4189..4208)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4399..4420
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4574..4593)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4693..4715
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4753..4771)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4839..4943
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4944..5012,5013..5804)
                     /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(5162..5181)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5975..6563
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6464..6483
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt
      181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
      241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
      481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
      541 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gatccataac ttcgtatagg
     1441 atactttata cgaagttatg tagctggagc aatttgaaaa gattgttgta ttacctctag
     1501 ctgcgcttaa taacttcgta tagtatacct tatacgaagt tatactagtc ctcctgtact
     1561 cacctgggtc acggtcacca cgccgccgtc ctcgaagttc atcacgcgct cccacttgaa
     1621 gccctcgggg aaggacagct tcaagtagtc ggggatgtcg gcggggtgct tcacgtaggc
     1681 cttggagccg tacatgaact gaggggacag gatgtcccag gcgaagggca gggggccacc
     1741 cttggtcacc ttcagcttgg cggtctgggt gccctcgtag gggcggccct cgccctcgcc
     1801 ctcgatctcg aactcgtggc cgttcacgga gccctccatg tgcaccttga agcgcatgaa
     1861 ctccttgatg atggccatgt tatcctcctc gcccttgctc accatggtgg ctagcataac
     1921 ttcgtataaa gtatcctata cgaagttatt tgccttaacc cagaaattat cactgttatt
     1981 ctttagatag gtgcaaagaa taacttcgta taaggtatac tatacgaagt tatggtaccg
     2041 aagttcctat tcttcaaata gtataggaac ttcaattgtt attggtgcaa agaatcactt
     2101 ccttaagtct ttatactgcc caggaagttc ctattcttca aaaggtatag gaacttcggc
     2161 gcgccttact tgtacagctc gtccatgccg ccggtggagt ggcggccctc ggcgcgttcg
     2221 tactgttcca cgatggtgta gtcctcgttg tgggaggtga tgtccaactt gatgttgacg
     2281 ttgtaggcgc cgggcagctg cacgggcttc ttggccttgt aggtggtctt gacctcagcg
     2341 tcgtagtggc cgccgtcctt cagcttcagc ctctgcttga tctcgccctt cagggcgccg
     2401 tcctcggggt acatccgctc ggaggaggcc tcccagccca tggtcttctt ctgcattacg
     2461 gggccgtcgg aggggaagtt ggtgccgcgc agcttcacct tgtagatgaa ctcgccgtcc
     2521 tgcagggagg agtcctgtgg gacgacatga cttaaccagt taaccatgtt ctctgaacat
     2581 gagaagggct ggccactctc cagagctcga agttcctata ctatttgaag aataggaact
     2641 tcttgcctta acccagaaat tatcactgtt attctttaga atggtgcaaa gagaagttcc
     2701 tatacctttt gaagaatagg aacttcgaat tcgatatcaa gcttatcgat aatcaacctc
     2761 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
     2821 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
     2881 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
     2941 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
     3001 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
     3061 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
     3121 acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctatgttg
     3181 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
     3241 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
     3301 ctcagacgag tcggatctcc ctttgggccg cctccccgca tcgataccga gcgctgctcg
     3361 agagatctac gggtggcatc cctgtgaccc ctccccagtg cctctcctgg ccctggaagt
     3421 tgccactcca gtgcccacca gccttgtcct aataaaatta agttgcatca ttttgtctga
     3481 ctaggtgtcc ttctataata ttatggggtg gaggggggtg gtatggagca aggggcaagt
     3541 tgggaagaca acctgtaggg cctgcggggt ctattgggaa ccaagctgga gtgcagtggc
     3601 acaatcttgg ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc tgcctcagcc
     3661 tcccgagttg ttgggattcc aggcatgcat gaccaggctc agctaatttt tgtttttttg
     3721 gtagagacgg ggtttcacca tattggccag gctggtctcc aactcctaat ctcaggtgat
     3781 ctacccacct tggcctccca aattgctggg attacaggcg tgaaccactg ctcccttccc
     3841 tgtccttctg attttgtagg taaccacgtg cggaccgagc ggccgcagga acccctagtg
     3901 atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag
     3961 gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc
     4021 ctgcaggggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     4081 atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     4141 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     4201 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     4261 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgatttgg
     4321 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     4381 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     4441 cgggctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     4501 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaattttat
     4561 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc
     4621 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     4681 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     4741 cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg
     4801 tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     4861 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     4921 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     4981 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     5041 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     5101 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     5161 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     5221 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     5281 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     5341 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     5401 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     5461 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     5521 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     5581 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     5641 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     5701 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     5761 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     5821 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     5881 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     5941 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     6001 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     6061 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     6121 tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     6181 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     6241 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     6301 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     6361 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     6421 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     6481 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     6541 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     6601 tttgctggcc ttttgctcac atgt
//
