LOCUS       sequence-271929-        6614 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 6614)
  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 6614)
  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..6614
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     27..46
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   188..317
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        418..1596
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          649..1587
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1544..1564
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1603..1636)
                     /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(1687..1720)
                     /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     1919..1937
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      2079..2088
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    2093..2126
                     /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    2177..2210
                     /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    2217..2250
                     /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(2621..2640)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(2786..2819)
                     /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    2928..3516
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2981..3001)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3399..3410)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3548..4024
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3747..3766)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4064..4204
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4064..4193
                     /label=AAV2 ITR
     rep_origin      4279..4734
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4366..4385)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4576..4597
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4751..4770)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4870..4892
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4930..4948)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5016..5120
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5121..5189,5190..5981)
                     /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(5339..5358)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(6152..6614,1..126)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
       61 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
      121 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
      181 cacatgtcct gcaggcagct gcgcgctcgc tcgctcactg aggccgcccg ggcgtcgggc
      241 gacctttggt cgcccggcct cagtgagcga gcgagcgcgc agagagggag tggccaactc
      301 catcactagg ggttcctgcg gccgcacgcg taagctttgc aaagatggat aaagttttaa
      361 acagagagga atctttgcag ctaatggacc ttctaggtct tgaaaggagt gggaattggc
      421 tccggtgccc gtcagtgggc agagcgcaca tcgcccacag tccccgagaa gttgggggga
      481 ggggtcggca attgaaccgg tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat
      541 gtcgtgtact ggctccgcct ttttcccgag ggtgggggag aaccgtatat aagtgcagta
      601 gtcgccgtga acgttctttt tcgcaacggg tttgccgcca gaacacaggt aagtgccgtg
      661 tgtggttccc gcgggcctgg cctctttacg ggttatggcc cttgcgtgcc ttgaattact
      721 tccacctggc tgcagtacgt gattcttgat cccgagcttc gggttggaag tgggtgggag
      781 agttcgaggc cttgcgctta aggagcccct tcgcctcgtg cttgagttga ggcctggcct
      841 gggcgctggg gccgccgcgt gcgaatctgg tggcaccttc gcgcctgtct cgctgctttc
      901 gataagtctc tagccattta aaatttttga tgacctgctg cgacgctttt tttctggcaa
      961 gatagtcttg taaatgcggg ccaagatctg cacactggta tttcggtttt tggggccgcg
     1021 ggcggcgacg gggcccgtgc gtcccagcgc acatgttcgg cgaggcgggg cctgcgagcg
     1081 cggccaccga gaatcggacg ggggtagtct caagctggcc ggcctgctct ggtgcctggc
     1141 ctcgcgccgc cgtgtatcgc cccgccctgg gcggcaaggc tggcccggtc ggcaccagtt
     1201 gcgtgagcgg aaagatggcc gcttcccggc cctgctgcag ggagctcaaa atggaggacg
     1261 cggcgctcgg gagagcgggc gggtgagtca cccacacaaa ggaaaagggc ctttccgtcc
     1321 tcagccgtcg cttcatgtga ctccacggag taccgggcgc cgtccaggca cctcgattag
     1381 ttctcgagct tttggagtac gtcgtcttta ggttgggggg aggggtttta tgcgatggag
     1441 tttccccaca ctgagtgggt ggagactgaa gttaggccag cttggcactt gatgtaattc
     1501 tccttggaat ttgccctttt tgagtttgga tcttggttca ttctcaagcc tcagacagtg
     1561 gttcaaagtt tttttcttcc atttcaggtg tcgtgaggat ccataacttc gtataggata
     1621 ctttatacga agttatgtag ctggagcaat ttgaaaagat tgttgtatta cctctagctg
     1681 cgcttaataa cttcgtatag tataccttat acgaagttat actagtcctc ctgtactcac
     1741 ctgggtcacg gtcaccacgc cgccgtcctc gaagttcatc acgcgctccc acttgaagcc
     1801 ctcggggaag gacagcttca agtagtcggg gatgtcggcg gggtgcttca cgtaggcctt
     1861 ggagccgtac atgaactgag gggacaggat gtcccaggcg aagggcaggg ggccaccctt
     1921 ggtcaccttc agcttggcgg tctgggtgcc ctcgtagggg cggccctcgc cctcgccctc
     1981 gatctcgaac tcgtggccgt tcacggagcc ctccatgtgc accttgaagc gcatgaactc
     2041 cttgatgatg gccatgttat cctcctcgcc cttgctcacc atggtggcta gcataacttc
     2101 gtataaagta tcctatacga agttatttgc cttaacccag aaattatcac tgttattctt
     2161 tagataggtg caaagaataa cttcgtataa ggtatactat acgaagttat ggtaccgaag
     2221 ttcctattct tcaaatagta taggaacttc aattgttatt ggtgcaaaga atcacttcct
     2281 taagtcttta tactgcccag gaagttccta ttcttcaaaa ggtataggaa cttcggcgcg
     2341 ccttacttgt acagctcgtc catgccgccg gtggagtggc ggccctcggc gcgttcgtac
     2401 tgttccacga tggtgtagtc ctcgttgtgg gaggtgatgt ccaacttgat gttgacgttg
     2461 taggcgccgg gcagctgcac gggcttcttg gccttgtagg tggtcttgac ctcagcgtcg
     2521 tagtggccgc cgtccttcag cttcagcctc tgcttgatct cgcccttcag ggcgccgtcc
     2581 tcggggtaca tccgctcgga ggaggcctcc cagcccatgg tcttcttctg cattacgggg
     2641 ccgtcggagg ggaagttggt gccgcgcagc ttcaccttgt agatgaactc gccgtcctgc
     2701 agggaggagt cctgtgggac gacatgactt aaccagttaa ccatgttctc tgaacatgag
     2761 aagggctggc cactctccag agctcgaagt tcctatacta tttgaagaat aggaacttct
     2821 tgccttaacc cagaaattat cactgttatt ctttagaatg gtgcaaagag aagttcctat
     2881 accttttgaa gaataggaac ttcgaattcg atatcaagct tatcgataat caacctctgg
     2941 attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat
     3001 gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt
     3061 tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca
     3121 ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg
     3181 ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg
     3241 aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca
     3301 attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tatgttgcca
     3361 cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc
     3421 ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc
     3481 agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgagcg ctgctcgaga
     3541 gatctacggg tggcatccct gtgacccctc cccagtgcct ctcctggccc tggaagttgc
     3601 cactccagtg cccaccagcc ttgtcctaat aaaattaagt tgcatcattt tgtctgacta
     3661 ggtgtccttc tataatatta tggggtggag gggggtggta tggagcaagg ggcaagttgg
     3721 gaagacaacc tgtagggcct gcggggtcta ttgggaacca agctggagtg cagtggcaca
     3781 atcttggctc actgcaatct ccgcctcctg ggttcaagcg attctcctgc ctcagcctcc
     3841 cgagttgttg ggattccagg catgcatgac caggctcagc taatttttgt ttttttggta
     3901 gagacggggt ttcaccatat tggccaggct ggtctccaac tcctaatctc aggtgatcta
     3961 cccaccttgg cctcccaaat tgctgggatt acaggcgtga accactgctc ccttccctgt
     4021 ccttctgatt ttgtaggtaa ccacgtgcgg accgagcggc cgcaggaacc cctagtgatg
     4081 gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc
     4141 gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg
     4201 caggggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     4261 cgtcaaagca accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     4321 ttacgcgcag cgtgaccgct acacttgcca gcgccttagc gcccgctcct ttcgctttct
     4381 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     4441 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg
     4501 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     4561 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac tctatctcgg
     4621 gctattcttt tgatttataa gggattttgc cgatttcggt ctattggtta aaaaatgagc
     4681 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt
     4741 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa
     4801 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     4861 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     4921 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     4981 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     5041 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     5101 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     5161 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     5221 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     5281 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     5341 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     5401 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     5461 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     5521 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     5581 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     5641 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     5701 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     5761 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     5821 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     5881 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     5941 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     6001 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     6061 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     6121 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     6181 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     6241 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     6301 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     6361 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     6421 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     6481 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     6541 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     6601 gcagggtcgg aaca
//
