LOCUS Exported 6616 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Intersectional viral expression of mCherry in cells expressing Flp AND NOT Cre. ACCESSION . VERSION . KEYWORDS pAAV-Ef1a-Coff/Fon-mCherry SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6616) 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 6616) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6616 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..130 /label=AAV2 ITR (alternate) /note="Functional equivalent of wild-type AAV2 ITR" promoter 231..1409 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 462..1400 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 1357..1377 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" protein_bind 1416..1449 /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." protein_bind 1500..1533 /label=loxN /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GTATACCT). loxN sites are compatible with each other, but incompatible with loxP or lox2272 sites." regulatory 1540..1549 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" primer_bind complement(1691..1709) /label=mCherry-R /note="mCherry, reverse primer" protein_bind complement(1908..1941) /label=lox2272 /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GGATACTT). lox2272 sites are compatible with each other, but incompatible with loxP or loxN sites (Lee and Saito, 1988)." protein_bind complement(1992..2025) /label=loxN /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GTATACCT). loxN sites are compatible with each other, but incompatible with loxP or lox2272 sites." primer_bind complement(2436..2455) /label=mCherry-F /note="mCherry, forward primer" misc_feature 2743..3331 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2796..2816) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3214..3225) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3363..3839 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(3562..3581) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 3879..4019 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 3879..4008 /label=AAV2 ITR rep_origin 4094..4549 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4181..4200) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4391..4412 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4566..4585) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4685..4707 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4745..4763) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4831..4935 /gene="bla" /label=AmpR promoter CDS 4936..5796 /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(5154..5173) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5967..6555 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6456..6475 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt 61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact 121 aggggttcct gcggccgcac gcgtaagctt tgcaaagatg gataaagttt taaacagaga 181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg 241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg 301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt 361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg 421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt 481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct 541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga 601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct 661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt 721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc 781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg 841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac 901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc 961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag 1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct 1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg 1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga 1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc 1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg 1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa 1381 gtttttttct tccatttcag gtgtcgtgag gatccataac ttcgtatagg atactttata 1441 cgaagttatg tagctggagc aatttgaaaa gattgttgta ttacctctag ctgcgcttaa 1501 taacttcgta tagtatacct tatacgaagt tatactagtg ccaccatggt gagcaagggc 1561 gaggaggata acatggccat catcaaggag ttcatgcgct tcaaggtgca catggagggc 1621 tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc 1681 acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc 1741 ctgtcccctc agttcatgta cggctccaag gcctacgtga agcaccccgc cgacatcccc 1801 gactacttga agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag 1861 gacggcggcg tggtgaccgt gacccaggtg agtacaggag ggctagcata acttcgtata 1921 aagtatccta tacgaagtta tttgccttaa cccagaaatt atcactgtta ttctttagat 1981 aggtgcaaag aataacttcg tataaggtat actatacgaa gttatggtac cgaagttcct 2041 attcttcaaa tagtatagga acttcaattg ttattggtgc aaagaatcac ttccttaagt 2101 ctttatactg cccaggaagt tcctattctt caaaaggtat aggaacttcg gcgcgcctta 2161 cttgtacagc tcgtccatgc cgccggtgga gtggcggccc tcggcgcgtt cgtactgttc 2221 cacgatggtg tagtcctcgt tgtgggaggt gatgtccaac ttgatgttga cgttgtaggc 2281 gccgggcagc tgcacgggct tcttggcctt gtaggtggtc ttgacctcag cgtcgtagtg 2341 gccgccgtcc ttcagcttca gcctctgctt gatctcgccc ttcagggcgc cgtcctcggg 2401 gtacatccgc tcggaggagg cctcccagcc catggtcttc ttctgcatta cggggccgtc 2461 ggaggggaag ttggtgccgc gcagcttcac cttgtagatg aactcgccgt cctgcaggga 2521 ggagtcctgt gggacgacat gacttaacca gttaaccatg ttctctgaac atgagaaggg 2581 ctggccactc tccagagctc gaagttccta tactatttga agaataggaa cttcttgcct 2641 taacccagaa attatcactg ttattcttta gaatggtgca aagagaagtt cctatacctt 2701 ttgaagaata ggaacttcga attcgatatc aagcttatcg ataatcaacc tctggattac 2761 aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga 2821 tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc 2881 tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa 2941 cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc 3001 acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc 3061 atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc 3121 gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctatgt tgccacctgg 3181 attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct 3241 tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg 3301 agtcggatct ccctttgggc cgcctccccg catcgatacc gagcgctgct cgagagatct 3361 acgggtggca tccctgtgac ccctccccag tgcctctcct ggccctggaa gttgccactc 3421 cagtgcccac cagccttgtc ctaataaaat taagttgcat cattttgtct gactaggtgt 3481 ccttctataa tattatgggg tggagggggg tggtatggag caaggggcaa gttgggaaga 3541 caacctgtag ggcctgcggg gtctattggg aaccaagctg gagtgcagtg gcacaatctt 3601 ggctcactgc aatctccgcc tcctgggttc aagcgattct cctgcctcag cctcccgagt 3661 tgttgggatt ccaggcatgc atgaccaggc tcagctaatt tttgtttttt tggtagagac 3721 ggggtttcac catattggcc aggctggtct ccaactccta atctcaggtg atctacccac 3781 cttggcctcc caaattgctg ggattacagg cgtgaaccac tgctcccttc cctgtccttc 3841 tgattttgta ggtaaccacg tgcggaccga gcggccgcag gaacccctag tgatggagtt 3901 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg 3961 acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg 4021 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca 4081 aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg 4141 cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct 4201 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta 4261 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt 4321 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg 4381 ttctttaata gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat 4441 tcttttgatt tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt 4501 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact 4561 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc 4621 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc 4681 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga 4741 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag 4801 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa 4861 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat 4921 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg 4981 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa 5041 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt 5101 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt 5161 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat 5221 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg 5281 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta 5341 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat 5401 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag 5461 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa 5521 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca 5581 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc 5641 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt 5701 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc 5761 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat 5821 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt 5881 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac 5941 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc 6001 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca 6061 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta 6121 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct 6181 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg 6241 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc 6301 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta 6361 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg 6421 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt 6481 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg 6541 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg 6601 ccttttgctc acatgt //