LOCUS Exported 6612 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Intersectional viral expression of oScarlet in cells expressing both Cre AND Flp. ACCESSION . VERSION . KEYWORDS pAAV-Ef1a-Con/Fon-oScarlet SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6612) 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 6612) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6612 /organism="synthetic DNA construct" /mol_type="other DNA" 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 1426..1459 /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 1510..1543 /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 1624..1642 /label=mCherry-R /note="mCherry, reverse primer" regulatory 1772..1781 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" protein_bind complement(1786..1819) /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(1870..1903) /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(2314..2333) /label=mCherry-F /note="mCherry, forward primer" misc_feature 2739..3327 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2792..2812) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3210..3221) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3359..3835 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(3558..3577) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 3875..4015 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 3875..4004 /label=AAV2 ITR rep_origin 4090..4545 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4177..4196) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4387..4408 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4562..4581) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4681..4703 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4741..4759) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4827..4931 /gene="bla" /label=AmpR promoter CDS 4932..5792 /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(5150..5169) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5963..6551 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6452..6471 /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 tatgtcgacc ctcctgtact 1561 cacctggagc cgtacatgaa ctgaggggac aggatgtccc aggagaaggg cagggggcca 1621 cccttggtca ccttcagctt ggcggtctgg gtgccctcgt aggggcggcc ctcgccctcg 1681 ccctcgatct cgaactcgtg gccgttcatg gagccctcca tgtgcacctt gaaccgcatg 1741 aactccttga tcactgcctc gcccttgctc accatggtgg ctagcataac ttcgtataaa 1801 gtatcctata cgaagttatt tgccttaacc cagaaattat cactgttatt ctttagatag 1861 gtgcaaagaa taacttcgta taaggtatac tatacgaagt tatggtaccg aagttcctat 1921 tcttcaaata gtataggaac ttcaattgtt attggtgcaa agaatcactt ccttaagtct 1981 ttatactgcc caggaagttc ctattcttca aaaggtatag gaacttcggc gcgccttact 2041 tgtacagctc gtccatgccg ccggtggagt ggcggccctc ggagcgttcg tactgttcca 2101 ccacggtgta gtcctcgttg tgggaggtga tgtccaactt gcggtccacg ttgtaggcgc 2161 cgggcatctg cacgggcttc ttggccttgt aggtggtctt gaagtccgcc aggtagcgcc 2221 cgccgtcctt caggcgcagg gccatcttaa tgtcgccctt cagcacgccg tcctcggggt 2281 acaaccgctc ggtggacgct tcccagccca ttgtcttctt ctgcattacg gggccgtcag 2341 gagggaagtt ggtgccgcgg agcttcacct tgtagatcag ggtgccgtcc tccagggagg 2401 tgtcctgggt cacggtcacg gcgccgccgt cctcgaagtt catcacgcgg tcccacttga 2461 agccctcggg gaaggactgc ttatagtagt cggggatgtc ggcggggtgc ttggtgaagg 2521 ccctgtggga cgacatgact taaccagtta accatgttct ctgaacatga gaagggctgg 2581 ccactctcca tctagagaag ttcctatact atttgaagaa taggaacttc ttgccttaac 2641 ccagaaatta tcactgttat tctttagaat agtgcaaaga gaagttccta taccttttga 2701 agaataggaa cttcgaattc gatatcaagc ttatcgataa tcaacctctg gattacaaaa 2761 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg 2821 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct 2881 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg 2941 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct 3001 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg 3061 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg 3121 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctatgttgcc acctggattc 3181 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc 3241 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc 3301 ggatctccct ttgggccgcc tccccgcatc gataccgagc gctgctcgag agatctacgg 3361 gtggcatccc tgtgacccct ccccagtgcc tctcctggcc ctggaagttg ccactccagt 3421 gcccaccagc cttgtcctaa taaaattaag ttgcatcatt ttgtctgact aggtgtcctt 3481 ctataatatt atggggtgga ggggggtggt atggagcaag gggcaagttg ggaagacaac 3541 ctgtagggcc tgcggggtct attgggaacc aagctggagt gcagtggcac aatcttggct 3601 cactgcaatc tccgcctcct gggttcaagc gattctcctg cctcagcctc ccgagttgtt 3661 gggattccag gcatgcatga ccaggctcag ctaatttttg tttttttggt agagacgggg 3721 tttcaccata ttggccaggc tggtctccaa ctcctaatct caggtgatct acccaccttg 3781 gcctcccaaa ttgctgggat tacaggcgtg aaccactgct cccttccctg tccttctgat 3841 tttgtaggta accacgtgcg gaccgagcgg ccgcaggaac ccctagtgat ggagttggcc 3901 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc 3961 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc 4021 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc 4081 aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca 4141 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct 4201 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt 4261 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac 4321 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct 4381 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg ggctattctt 4441 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac 4501 aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca 4561 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg 4621 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct 4681 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg 4741 gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt 4801 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac 4861 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa 4921 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat 4981 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc 5041 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga 5101 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg 5161 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc 5221 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag 5281 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc 5341 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg 5401 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg 5461 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac 5521 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac 5581 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg 5641 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg 5701 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg 5761 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac 5821 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg 5881 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg 5941 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc 6001 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc 6061 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt 6121 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc 6181 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact 6241 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac 6301 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag 6361 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg 6421 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg 6481 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga 6541 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt 6601 ttgctcacat gt //