LOCUS Exported 6614 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Intersectional viral expression of oScarlet in cells expressing Cre AND NOT Flp. ACCESSION . VERSION . KEYWORDS pAAV-Ef1a-Con/Foff 2.0-oScarlet SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6614) 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 6614) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6614 /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(1788..1821) /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(1872..1905) /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." protein_bind 1912..1945 /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" primer_bind 2295..2314 /label=mCherry-F /note="mCherry, forward primer" protein_bind complement(2599..2632) /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" misc_feature 2741..3329 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2794..2814) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3212..3223) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3361..3837 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(3560..3579) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 3877..4017 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 3877..4006 /label=AAV2 ITR rep_origin 4092..4547 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4179..4198) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4389..4410 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4564..4583) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4683..4705 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4743..4761) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4829..4933 /gene="bla" /label=AmpR promoter CDS 4934..5794 /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(5152..5171) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5965..6553 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6454..6473 /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 cactagtata acttcgtata 1801 aagtatccta tacgaagtta tttgccttaa cccagaaatt atcactgtta ttctttagat 1861 aggtgcaaag aataacttcg tataaggtat actatacgaa gttatggtac cgaagttcct 1921 attctctaga aagtatagga acttcgcaga atggtagctg gattgtagct gctattagca 1981 atatgaaacc tcttagaagt tcctattctt caaaaggtat aggaacttcg gcgcgcctgg 2041 agagtggcca gcccttctca tgttcagaga acatggttaa ctggttaagt catgtcgtcc 2101 cacagggcct tcaccaagca ccccgccgac atccccgact actataagca gtccttcccc 2161 gagggcttca agtgggaccg cgtgatgaac ttcgaggacg gcggcgccgt gaccgtgacc 2221 caggacacct ccctggagga cggcaccctg atctacaagg tgaagctccg cggcaccaac 2281 ttccctcctg acggccccgt aatgcagaag aagacaatgg gctgggaagc gtccaccgag 2341 cggttgtacc ccgaggacgg cgtgctgaag ggcgacatta agatggccct gcgcctgaag 2401 gacggcgggc gctacctggc ggacttcaag accacctaca aggccaagaa gcccgtgcag 2461 atgcccggcg cctacaacgt ggaccgcaag ttggacatca cctcccacaa cgaggactac 2521 accgtggtgg aacagtacga acgctccgag ggccgccact ccaccggcgg catggacgag 2581 ctgtacaagt aagctagcga agttcctata ctttctagag aataggaact tctaagcaga 2641 caatcttgta atacatcagc agctagtaga ttgctccagc tagaagttcc tatacctttt 2701 gaagaatagg aacttcgaat tcgatatcaa gcttatcgat aatcaacctc tggattacaa 2761 aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata 2821 cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc 2881 cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg 2941 tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac 3001 ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat 3061 cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt 3121 ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctatgttg ccacctggat 3181 tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc 3241 ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag 3301 tcggatctcc ctttgggccg cctccccgca tcgataccga gcgctgctcg agagatctac 3361 gggtggcatc cctgtgaccc ctccccagtg cctctcctgg ccctggaagt tgccactcca 3421 gtgcccacca gccttgtcct aataaaatta agttgcatca ttttgtctga ctaggtgtcc 3481 ttctataata ttatggggtg gaggggggtg gtatggagca aggggcaagt tgggaagaca 3541 acctgtaggg cctgcggggt ctattgggaa ccaagctgga gtgcagtggc acaatcttgg 3601 ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc tgcctcagcc tcccgagttg 3661 ttgggattcc aggcatgcat gaccaggctc agctaatttt tgtttttttg gtagagacgg 3721 ggtttcacca tattggccag gctggtctcc aactcctaat ctcaggtgat ctacccacct 3781 tggcctccca aattgctggg attacaggcg tgaaccactg ctcccttccc tgtccttctg 3841 attttgtagg taaccacgtg cggaccgagc ggccgcagga acccctagtg atggagttgg 3901 ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac 3961 gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc 4021 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa 4081 gcaaccatag tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg 4141 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc 4201 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg 4261 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc 4321 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt 4381 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cgggctattc 4441 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta 4501 acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaattttat ggtgcactct 4561 cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc 4621 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt 4681 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa 4741 gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac 4801 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat 4861 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg 4921 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc 4981 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga 5041 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga 5101 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg 5161 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc 5221 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac 5281 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact 5341 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca 5401 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg 5461 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact 5521 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg 5581 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg 5641 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat 5701 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc 5761 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat 5821 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt 5881 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc 5941 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt 6001 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac 6061 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt 6121 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct 6181 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga 6241 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac 6301 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg 6361 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt 6421 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc 6481 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg 6541 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc 6601 ttttgctcac atgt //