LOCUS       sequence-265123-        7109 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Intersectional viral expression of ChRmine-p2a-oScarlet in cells 
            expressing Flp AND NOT Cre.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7109)
  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 7109)
  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-nEF-Coff/Fon-ChRmine-oScarlet
FEATURES             Location/Qualifiers
     source          1..7109
                     /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        188..399
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             412..680
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     protein_bind    708..741
                     /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(1136..1155)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    complement(1419..1452)
                     /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(1503..1536)
                     /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     1617..1635
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(1769..1825)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     protein_bind    2608..2641
                     /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    2692..2725
                     /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      3078..3087
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(3094..3127)
                     /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    3236..3824
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3289..3309)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3707..3718)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3856..4332
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4055..4074)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4372..4512
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4372..4501
                     /label=AAV2 ITR
     rep_origin      4587..5042
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4674..4693)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4884..4905
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5059..5078)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5178..5200
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5238..5256)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5324..5428
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5429..5497,5498..6289)
                     /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(5647..5666)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6460..7048
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6949..6968
                     /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 cgcgtaagga tctgcgatcg ctccggtgcc
      181 cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc
      241 aattgaacgg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac
      301 tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg
      361 aacgttcttt ttcgcaacgg gtttgccgcc agaacacagc tgaagcttcg aggggctcgc
      421 atctctcctt cacgcgcccg ccgccctacc tgaggccgcc atccacgccg gttgagtcgc
      481 gttctgccgc ctcccgcctg tggtgcctcc tgaactgcgt ccgccgtcta ggtaagttta
      541 aagctcaggt cgagaccggg cctttgtccg gcgctccctt ggagcctacc tagactcagc
      601 cggctctcca cgctttgcct gaccctgctt gctcaactct acgtctttgt ttcgttttct
      661 gttctgcgcc gttacagatc caagctgtga ccggcgccta cggatccgaa gttcctattc
      721 ttcaaatagt ataggaactt cgcagagtag tagctggatt gtagctgcta ttagcaatct
      781 aaaacctctt agaagttcct attcttcaaa aggtatagga acttcggcgc gcctcattac
      841 acctcgttct cgtagcagaa cttgtacagc tcgtccatgc cgccggtgga gtggcggccc
      901 tcggagcgtt cgtactgttc caccacggtg tagtcctcgt tgtgggaggt gatgtccaac
      961 ttgcggtcca cgttgtaggc gccgggcatc tgcacgggct tcttggcctt gtaggtggtc
     1021 ttgaagtccg ccaggtagcg cccgccgtcc ttcaggcgca gggccatctt aatgtcgccc
     1081 ttcagcacgc cgtcctcggg gtacaaccgc tcggtggacg cttcccagcc cattgtcttc
     1141 ttctgcatta cggggccgtc aggagggaag ttggtgccgc ggagcttcac cttgtagatc
     1201 agggtgccgt cctccaggga ggtgtcctgg gtcacggtca cggcgccgcc gtcctcgaag
     1261 ttcatcacgc ggtcccactt gaagccctcg gggaaggact gcttatagta gtcggggatg
     1321 tcggcggggt gcttggtgaa ggccctgtgg gacgacatga cttaaccagt taaccatgtt
     1381 ctctgaacat gagaagggct ggccactctc cagtcgacat aacttcgtat aggatacttt
     1441 atacgaagtt atgcagaatg gtagctggat tgtagctgct attagcaata tgaaacctct
     1501 taataacttc gtatagtata ccttatacga agttatacta gtcctcctgt actcacctgg
     1561 agccgtacat gaactgaggg gacaggatgt cccaggagaa gggcaggggg ccacccttgg
     1621 tcaccttcag cttggcggtc tgggtgccct cgtaggggcg gccctcgccc tcgccctcga
     1681 tctcgaactc gtggccgttc atggagccct ccatgtgcac cttgaaccgc atgaactcct
     1741 tgatcactgc ctcgcccttg ctcaccatgg gaccggggtt ttcttccacg tctcctgctt
     1801 gctttaacag agagaagttc gtggctccgg agccgggcgc gcccacgttg atgtcgatct
     1861 ggtccagggg gatgtactcg ccctcgctgg tgatcctgct cttggcggcc gcagacagtc
     1921 tccgcaggcg ctcgccattg ctgctcagcc tcatatccag ctccagcttc tggaactcat
     1981 cctcgtcgtg tcccagctgc tccagcagcc gaaacagctc ctcatcatag gtctttctga
     2041 accgggccag ggcaaagccg taacaggact tggccacgat gtcgcacacg cagtgggcca
     2101 cctctgtcca gttatcgtcg atccagccga agcctctagg gcagatggcc cacaccaggg
     2161 gaaacagaat ccacagcatg gagaaggtaa acacggcgaa cttcagcacg tgcaggctgt
     2221 gctcggcgcc tgtatcctgg gccagctggg ccagccggct atactgcttg gccacgatgc
     2281 tcatcacgat agagtaggcg aagataaacc agaaacagcc aaagccatac caggcgtagg
     2341 cgccattcct cagcctaggg tcgccctctg cgtagaactc ggccagcacg ccagacatca
     2401 ggatggcaaa gatgatggcg ctacaagaca cgcgataggg ggccctcagc tgatacagca
     2461 gatcgtacac cagcattggg caggtcagca tgtagtcggc atacctaaac acatgagaaa
     2521 tagagtcata agcacatggg tcacatacag gagatatgta tataacatta atacaatttt
     2581 ataaaaaaaa gggggggcac aggtaccata acttcgtata aagtatccta tacgaagtta
     2641 tttgccttaa cccagaaatt atcactgtta ttctttagaa tggtgcaaag aataacttcg
     2701 tataaggtat actatacgaa gttatgctag caaccccgac acttacctgt aattgccgaa
     2761 gccgatgatg aaggggccca catcgccgaa gttgtggccc agtgtgtcgg ccacgatctt
     2821 gccgaagtag ctcagcatgg caaagaaatt cacaaacagg gccaggttct gctcctcctg
     2881 atagccggcg taaaaggagg ttctcttgct agggccccga gatgtccagg actcatagcc
     2941 caggtagaat gtggtgattg tcagcacgat aaagcaagac cagtgggcgc cgatggccac
     3001 ggagttcagc ttggtatcca gataccagcc gtccattgtg cccacgtaga acatctggtc
     3061 ggtgcctggt gcgtgtgcca tggtggcgag ctcgaagttc ctatactatt tgaagaatag
     3121 gaacttcttg ccttaaccca gaaattatca ctgttattct ttagaatggt gcaaagagaa
     3181 gttcctatac cttttgaaga ataggaactt cgaattcgat atcaagctta tcgataatca
     3241 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     3301 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     3361 tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc
     3421 cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg
     3481 gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc
     3541 cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg
     3601 cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgccta
     3661 tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc
     3721 agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct
     3781 tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat accgagcgct
     3841 gctcgagaga tctacgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg
     3901 gaagttgcca ctccagtgcc caccagcctt gtcctaataa aattaagttg catcattttg
     3961 tctgactagg tgtccttcta taatattatg gggtggaggg gggtggtatg gagcaagggg
     4021 caagttggga agacaacctg tagggcctgc ggggtctatt gggaaccaag ctggagtgca
     4081 gtggcacaat cttggctcac tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct
     4141 cagcctcccg agttgttggg attccaggca tgcatgacca ggctcagcta atttttgttt
     4201 ttttggtaga gacggggttt caccatattg gccaggctgg tctccaactc ctaatctcag
     4261 gtgatctacc caccttggcc tcccaaattg ctgggattac aggcgtgaac cactgctccc
     4321 ttccctgtcc ttctgatttt gtaggtaacc acgtgcggac cgagcggccg caggaacccc
     4381 tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac
     4441 caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca
     4501 gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     4561 accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg
     4621 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     4681 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     4741 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     4801 tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     4861 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     4921 tatctcgggc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     4981 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat
     5041 tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca
     5101 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     5161 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     5221 acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     5281 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     5341 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     5401 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     5461 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     5521 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     5581 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     5641 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     5701 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     5761 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     5821 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     5881 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     5941 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     6001 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     6061 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     6121 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     6181 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     6241 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     6301 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     6361 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     6421 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     6481 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     6541 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     6601 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     6661 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     6721 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     6781 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     6841 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     6901 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     6961 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     7021 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     7081 ggccttttgc tggccttttg ctcacatgt
//
