LOCUS       sequence-265094-        7169 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Intersectional viral expression of sRGECO in cells expressing Flp 
            AND NOT Cre.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7169)
  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 7169)
  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-sRGECO
FEATURES             Location/Qualifiers
     source          1..7169
                     /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."
     regulatory      1550..1559
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1571..1627
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKAGHAVRAIGRLSS"
     protein_bind    1922..1955
                     /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    2006..2039
                     /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    2046..2079
                     /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(2639..2658)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(3154..3187)
                     /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    3296..3884
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3349..3369)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3767..3778)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3916..4392
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4115..4134)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4432..4572
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4432..4561
                     /label=AAV2 ITR
     rep_origin      4647..5102
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4734..4753)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4944..4965
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5119..5138)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5238..5260
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5298..5316)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5384..5488
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5489..5557,5558..6349)
                     /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(5707..5726)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6520..7108
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7009..7028
                     /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 tatgtcgacg ccaccatggt
     1561 ggactcatca cgtcgtaagt ggaataaggc aggtcacgca gtcagagcta taggtcggct
     1621 gagctcaccc gtggtttccg agcggatgta ccccgaggac ggcgccctga agagcgagat
     1681 caagaagggg ctgaggctga aggacggcgg ccactacgcc gccgaggtca agaccaccta
     1741 caaggccaag aagcccgtgc agctgcccgg cgcctacatc gtggacatca agttggacat
     1801 cgtgtcccac aacgaggact acaccatcgt ggaacagtgc gaacgcgccg agggccgcca
     1861 ctccaccggc ggcatggacg agctgtacaa gggaggtaca ggtgagtaca ggagggctag
     1921 cataacttcg tataaagtat cctatacgaa gttatttgcc ttaacccaga aattatcact
     1981 gttattcttt agataggtgc aaagaataac ttcgtataag gtatactata cgaagttatg
     2041 gtaccgaagt tcctattctt caaatagtat aggaacttca attgttattg gtgcaaagaa
     2101 tcacttcctt aagtctttat actgcccagg aagttcctat tcttcaaaag gtataggaac
     2161 ttcggcgcgc cctacttcgc tgtcatcatt tgtacaaact cttcgtagtt tacctgacca
     2221 tccccatcga tgtctgctac cctgatcatt tcatcaacct cctcatctgt taacttctct
     2281 ccaaggtctg tcatcacgtg gcgaagctct gctgcgccga tgtagccatt gccgtcctta
     2341 tcaaacacgc ggaacgcttc gcgaatttcc tcttcactgt ctgtgtcatt catttttctt
     2401 gccatcatcg tcaggaactc agggaagtcg aatgtgccgt caccgtcggc atctacttca
     2461 ttgatcatgt cctgcagctc tgcttctgtg gggttctgcc ccagcgatcg gaacaccgtc
     2521 ccaagctcct tggttgttat cgtcccatcc ccgtccttgt caaataggga gaaagcctct
     2581 ttaaattctg cgatctgctc ttcagtcagg tcgtcacgcg tagcctccca gcccatggtc
     2641 ttcttctgca ttacggggcc gtcggggggg aagttggtgc cgcgcagctt caccttgtag
     2701 atgaatacgc cgtcctgcag ggaggagtcc tggttcacgt gaataatgcc gccgtcctcg
     2761 aagttcatca cgcggtccca cctgaagccc tcggggaagg acagcttgaa gtagtcgggg
     2821 atgtcggctg ggtgcttaat gtaggccttg gagccgtaca tgaactgagg ggacaggatg
     2881 tcccaggcga agggcagggg gccacccttg gtcaccttca gcttagcggt ctgaaaggcc
     2941 tcgtaggggc ggccctcgcc ctcgccctcg atctcgaact cgtggccgtt cacggagccc
     3001 tccatgtgca ccttgaagcg catgaactcc ttgatgatgg ccatgttatc ctcctcgccc
     3061 ttgctcacca gactcccgcc tgtgggacga catgacttaa ccagttaacc atgttctctg
     3121 aacatgagaa gggctggcca ctctccatct agagaagttc ctatactatt tgaagaatag
     3181 gaacttcttg ccttaaccca gaaattatca ctgttattct ttagaatagt gcaaagagaa
     3241 gttcctatac cttttgaaga ataggaactt cgaattcgat atcaagctta tcgataatca
     3301 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     3361 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     3421 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc
     3481 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg
     3541 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc
     3601 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     3661 cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgccta
     3721 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc
     3781 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct
     3841 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat accgagcgct
     3901 gctcgagaga tctacgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg
     3961 gaagttgcca ctccagtgcc caccagcctt gtcctaataa aattaagttg catcattttg
     4021 tctgactagg tgtccttcta taatattatg gggtggaggg gggtggtatg gagcaagggg
     4081 caagttggga agacaacctg tagggcctgc ggggtctatt gggaaccaag ctggagtgca
     4141 gtggcacaat cttggctcac tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct
     4201 cagcctcccg agttgttggg attccaggca tgcatgacca ggctcagcta atttttgttt
     4261 ttttggtaga gacggggttt caccatattg gccaggctgg tctccaactc ctaatctcag
     4321 gtgatctacc caccttggcc tcccaaattg ctgggattac aggcgtgaac cactgctccc
     4381 ttccctgtcc ttctgatttt gtaggtaacc acgtgcggac cgagcggccg caggaacccc
     4441 tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac
     4501 caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca
     4561 gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     4621 accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg
     4681 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     4741 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     4801 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     4861 tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     4921 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     4981 tatctcgggc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     5041 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat
     5101 tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca
     5161 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     5221 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     5281 acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     5341 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     5401 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     5461 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     5521 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     5581 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     5641 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     5701 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     5761 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     5821 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     5881 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     5941 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     6001 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     6061 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     6121 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     6181 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     6241 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     6301 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     6361 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     6421 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     6481 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     6541 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     6601 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     6661 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     6721 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     6781 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     6841 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     6901 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     6961 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     7021 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     7081 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     7141 ggccttttgc tggccttttg ctcacatgt
//
