LOCUS       sequence-318624-        7159 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 7159)
  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 7159)
  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..7159
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      172..627
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(259..278)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     469..490
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(644..663)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     763..785
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(823..841)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        909..1013
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1014..1082,1083..1874)
                     /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(1232..1251)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2045..2633
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2534..2553
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   2695..2824
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        2925..4103
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          3156..4094
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     4051..4071
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(4110..4143)
                     /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(4194..4227)
                     /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      4234..4243
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4255..4311
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKAGHAVRAIGRLSS"
     protein_bind    4606..4639
                     /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    4690..4723
                     /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    4730..4763
                     /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(5323..5342)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(5838..5871)
                     /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    5980..6568
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6033..6053)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6451..6462)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    6600..7076
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(6799..6818)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   join(7116..7159,1..97)
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   join(7116..7159,1..86)
                     /label=AAV2 ITR
ORIGIN
        1 cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg
       61 gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc
      121 cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct
      181 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
      241 ccagcgcctt agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
      301 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
      361 ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg ccatcgccct
      421 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
      481 tccaaactgg aacaacactc aactctatct cgggctattc ttttgattta taagggattt
      541 tgccgatttc ggtctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
      601 ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc tgctctgatg
      661 ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
      721 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
      781 agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat
      841 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
      901 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
      961 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     1021 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     1081 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     1141 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     1201 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     1261 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     1321 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     1381 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     1441 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     1501 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     1561 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     1621 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     1681 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     1741 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     1801 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     1861 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     1921 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     1981 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     2041 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     2101 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     2161 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     2221 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     2281 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     2341 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     2401 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     2461 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     2521 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     2581 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     2641 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgtcctgca
     2701 ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc gtcgggcgac ctttggtcgc
     2761 ccggcctcag tgagcgagcg agcgcgcaga gagggagtgg ccaactccat cactaggggt
     2821 tcctgcggcc gcacgcgtaa gctttgcaaa gatggataaa gttttaaaca gagaggaatc
     2881 tttgcagcta atggaccttc taggtcttga aaggagtggg aattggctcc ggtgcccgtc
     2941 agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt
     3001 gaaccggtgc ctagagaagg tggcgcgggg taaactggga aagtgatgtc gtgtactggc
     3061 tccgcctttt tcccgagggt gggggagaac cgtatataag tgcagtagtc gccgtgaacg
     3121 ttctttttcg caacgggttt gccgccagaa cacaggtaag tgccgtgtgt ggttcccgcg
     3181 ggcctggcct ctttacgggt tatggccctt gcgtgccttg aattacttcc acctggctgc
     3241 agtacgtgat tcttgatccc gagcttcggg ttggaagtgg gtgggagagt tcgaggcctt
     3301 gcgcttaagg agccccttcg cctcgtgctt gagttgaggc ctggcctggg cgctggggcc
     3361 gccgcgtgcg aatctggtgg caccttcgcg cctgtctcgc tgctttcgat aagtctctag
     3421 ccatttaaaa tttttgatga cctgctgcga cgcttttttt ctggcaagat agtcttgtaa
     3481 atgcgggcca agatctgcac actggtattt cggtttttgg ggccgcgggc ggcgacgggg
     3541 cccgtgcgtc ccagcgcaca tgttcggcga ggcggggcct gcgagcgcgg ccaccgagaa
     3601 tcggacgggg gtagtctcaa gctggccggc ctgctctggt gcctggcctc gcgccgccgt
     3661 gtatcgcccc gccctgggcg gcaaggctgg cccggtcggc accagttgcg tgagcggaaa
     3721 gatggccgct tcccggccct gctgcaggga gctcaaaatg gaggacgcgg cgctcgggag
     3781 agcgggcggg tgagtcaccc acacaaagga aaagggcctt tccgtcctca gccgtcgctt
     3841 catgtgactc cacggagtac cgggcgccgt ccaggcacct cgattagttc tcgagctttt
     3901 ggagtacgtc gtctttaggt tggggggagg ggttttatgc gatggagttt ccccacactg
     3961 agtgggtgga gactgaagtt aggccagctt ggcacttgat gtaattctcc ttggaatttg
     4021 ccctttttga gtttggatct tggttcattc tcaagcctca gacagtggtt caaagttttt
     4081 ttcttccatt tcaggtgtcg tgaggatcca taacttcgta taggatactt tatacgaagt
     4141 tatgtagctg gagcaatttg aaaagattgt tgtattacct ctagctgcgc ttaataactt
     4201 cgtatagtat accttatacg aagttatgtc gacgccacca tggtggactc atcacgtcgt
     4261 aagtggaata aggcaggtca cgcagtcaga gctataggtc ggctgagctc acccgtggtt
     4321 tccgagcgga tgtaccccga ggacggcgcc ctgaagagcg agatcaagaa ggggctgagg
     4381 ctgaaggacg gcggccacta cgccgccgag gtcaagacca cctacaaggc caagaagccc
     4441 gtgcagctgc ccggcgccta catcgtggac atcaagttgg acatcgtgtc ccacaacgag
     4501 gactacacca tcgtggaaca gtgcgaacgc gccgagggcc gccactccac cggcggcatg
     4561 gacgagctgt acaagggagg tacaggtgag tacaggaggg ctagcataac ttcgtataaa
     4621 gtatcctata cgaagttatt tgccttaacc cagaaattat cactgttatt ctttagatag
     4681 gtgcaaagaa taacttcgta taaggtatac tatacgaagt tatggtaccg aagttcctat
     4741 tcttcaaata gtataggaac ttcaattgtt attggtgcaa agaatcactt ccttaagtct
     4801 ttatactgcc caggaagttc ctattcttca aaaggtatag gaacttcggc gcgccctact
     4861 tcgctgtcat catttgtaca aactcttcgt agtttacctg accatcccca tcgatgtctg
     4921 ctaccctgat catttcatca acctcctcat ctgttaactt ctctccaagg tctgtcatca
     4981 cgtggcgaag ctctgctgcg ccgatgtagc cattgccgtc cttatcaaac acgcggaacg
     5041 cttcgcgaat ttcctcttca ctgtctgtgt cattcatttt tcttgccatc atcgtcagga
     5101 actcagggaa gtcgaatgtg ccgtcaccgt cggcatctac ttcattgatc atgtcctgca
     5161 gctctgcttc tgtggggttc tgccccagcg atcggaacac cgtcccaagc tccttggttg
     5221 ttatcgtccc atccccgtcc ttgtcaaata gggagaaagc ctctttaaat tctgcgatct
     5281 gctcttcagt caggtcgtca cgcgtagcct cccagcccat ggtcttcttc tgcattacgg
     5341 ggccgtcggg ggggaagttg gtgccgcgca gcttcacctt gtagatgaat acgccgtcct
     5401 gcagggagga gtcctggttc acgtgaataa tgccgccgtc ctcgaagttc atcacgcggt
     5461 cccacctgaa gccctcgggg aaggacagct tgaagtagtc ggggatgtcg gctgggtgct
     5521 taatgtaggc cttggagccg tacatgaact gaggggacag gatgtcccag gcgaagggca
     5581 gggggccacc cttggtcacc ttcagcttag cggtctgaaa ggcctcgtag gggcggccct
     5641 cgccctcgcc ctcgatctcg aactcgtggc cgttcacgga gccctccatg tgcaccttga
     5701 agcgcatgaa ctccttgatg atggccatgt tatcctcctc gcccttgctc accagactcc
     5761 cgcctgtggg acgacatgac ttaaccagtt aaccatgttc tctgaacatg agaagggctg
     5821 gccactctcc atctagagaa gttcctatac tatttgaaga ataggaactt cttgccttaa
     5881 cccagaaatt atcactgtta ttctttagaa tagtgcaaag agaagttcct ataccttttg
     5941 aagaatagga acttcgaatt cgatatcaag cttatcgata atcaacctct ggattacaaa
     6001 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     6061 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc
     6121 ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt
     6181 ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc
     6241 tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc
     6301 gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg
     6361 gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctatgttgc cacctggatt
     6421 ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc
     6481 cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt
     6541 cggatctccc tttgggccgc ctccccgcat cgataccgag cgctgctcga gagatctacg
     6601 ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc cctggaagtt gccactccag
     6661 tgcccaccag ccttgtccta ataaaattaa gttgcatcat tttgtctgac taggtgtcct
     6721 tctataatat tatggggtgg aggggggtgg tatggagcaa ggggcaagtt gggaagacaa
     6781 cctgtagggc ctgcggggtc tattgggaac caagctggag tgcagtggca caatcttggc
     6841 tcactgcaat ctccgcctcc tgggttcaag cgattctcct gcctcagcct cccgagttgt
     6901 tgggattcca ggcatgcatg accaggctca gctaattttt gtttttttgg tagagacggg
     6961 gtttcaccat attggccagg ctggtctcca actcctaatc tcaggtgatc tacccacctt
     7021 ggcctcccaa attgctggga ttacaggcgt gaaccactgc tcccttccct gtccttctga
     7081 ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa cccctagtga tggagttggc
     7141 cactccctct ctgcgcgct
//
