LOCUS Exported 7157 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Intersectional viral expression of sRGECO in cells expressing both Cre AND Flp. ACCESSION . VERSION . KEYWORDS pAAV-Ef1a-Con/Fon-sRGECO SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7157) 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 7157) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7157 /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." CDS complement(1828..1884) /codon_start=1 /product="calmodulin-binding peptide from avian smooth muscle myosin light chain kinase" /label=calmodulin-binding peptide /translation="RRKWNKAGHAVRAIGRLSS" regulatory 1896..1905 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" protein_bind complement(1910..1943) /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(1994..2027) /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(2627..2646) /label=mCherry-F /note="mCherry, forward primer" misc_feature 3284..3872 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(3337..3357) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3755..3766) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3904..4380 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(4103..4122) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" repeat_region 4420..4560 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 4420..4549 /label=AAV2 ITR rep_origin 4635..5090 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4722..4741) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4932..4953 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(5107..5126) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 5226..5248 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(5286..5304) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 5372..5476 /gene="bla" /label=AmpR promoter CDS 5477..6337 /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(5695..5714) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 6508..7096 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6997..7016 /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 tatgtcgacc ctcctgtact cacctgtacc 1561 tcccttgtac agctcgtcca tgccgccggt ggagtggcgg ccctcggcgc gttcgcactg 1621 ttccacgatg gtgtagtcct cgttgtggga cacgatgtcc aacttgatgt ccacgatgta 1681 ggcgccgggc agctgcacgg gcttcttggc cttgtaggtg gtcttgacct cggcggcgta 1741 gtggccgccg tccttcagcc tcagcccctt cttgatctcg ctcttcaggg cgccgtcctc 1801 ggggtacatc cgctcggaaa ccacgggtga gctcagccga cctatagctc tgactgcgtg 1861 acctgcctta ttccacttac gacgtgatga gtccaccatg gtggctagca taacttcgta 1921 taaagtatcc tatacgaagt tatttgcctt aacccagaaa ttatcactgt tattctttag 1981 ataggtgcaa agaataactt cgtataaggt atactatacg aagttatggt accgaagttc 2041 ctattcttca aatagtatag gaacttcaat tgttattggt gcaaagaatc acttccttaa 2101 gtctttatac tgcccaggaa gttcctattc ttcaaaaggt ataggaactt cggcgcgccc 2161 tacttcgctg tcatcatttg tacaaactct tcgtagttta cctgaccatc cccatcgatg 2221 tctgctaccc tgatcatttc atcaacctcc tcatctgtta acttctctcc aaggtctgtc 2281 atcacgtggc gaagctctgc tgcgccgatg tagccattgc cgtccttatc aaacacgcgg 2341 aacgcttcgc gaatttcctc ttcactgtct gtgtcattca tttttcttgc catcatcgtc 2401 aggaactcag ggaagtcgaa tgtgccgtca ccgtcggcat ctacttcatt gatcatgtcc 2461 tgcagctctg cttctgtggg gttctgcccc agcgatcgga acaccgtccc aagctccttg 2521 gttgttatcg tcccatcccc gtccttgtca aatagggaga aagcctcttt aaattctgcg 2581 atctgctctt cagtcaggtc gtcacgcgta gcctcccagc ccatggtctt cttctgcatt 2641 acggggccgt cgggggggaa gttggtgccg cgcagcttca ccttgtagat gaatacgccg 2701 tcctgcaggg aggagtcctg gttcacgtga ataatgccgc cgtcctcgaa gttcatcacg 2761 cggtcccacc tgaagccctc ggggaaggac agcttgaagt agtcggggat gtcggctggg 2821 tgcttaatgt aggccttgga gccgtacatg aactgagggg acaggatgtc ccaggcgaag 2881 ggcagggggc cacccttggt caccttcagc ttagcggtct gaaaggcctc gtaggggcgg 2941 ccctcgccct cgccctcgat ctcgaactcg tggccgttca cggagccctc catgtgcacc 3001 ttgaagcgca tgaactcctt gatgatggcc atgttatcct cctcgccctt gctcaccaga 3061 ctcccgcctg tgggacgaca tgacttaacc agttaaccat gttctctgaa catgagaagg 3121 gctggccact ctccatctag agaagttcct atactatttg aagaatagga acttcttgcc 3181 ttaacccaga aattatcact gttattcttt agaatagtgc aaagagaagt tcctatacct 3241 tttgaagaat aggaacttcg aattcgatat caagcttatc gataatcaac ctctggatta 3301 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg 3361 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc 3421 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca 3481 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac 3541 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact 3601 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc 3661 cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctatg ttgccacctg 3721 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc 3781 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac 3841 gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgagcgctgc tcgagagatc 3901 tacgggtggc atccctgtga cccctcccca gtgcctctcc tggccctgga agttgccact 3961 ccagtgccca ccagccttgt cctaataaaa ttaagttgca tcattttgtc tgactaggtg 4021 tccttctata atattatggg gtggaggggg gtggtatgga gcaaggggca agttgggaag 4081 acaacctgta gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt ggcacaatct 4141 tggctcactg caatctccgc ctcctgggtt caagcgattc tcctgcctca gcctcccgag 4201 ttgttgggat tccaggcatg catgaccagg ctcagctaat ttttgttttt ttggtagaga 4261 cggggtttca ccatattggc caggctggtc tccaactcct aatctcaggt gatctaccca 4321 ccttggcctc ccaaattgct gggattacag gcgtgaacca ctgctccctt ccctgtcctt 4381 ctgattttgt aggtaaccac gtgcggaccg agcggccgca ggaaccccta gtgatggagt 4441 tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc 4501 gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg 4561 ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc 4621 aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac 4681 gcgcagcgtg accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc 4741 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt 4801 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg 4861 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac 4921 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta 4981 ttcttttgat ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat 5041 ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac 5101 tctcagtaca atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc 5161 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac 5221 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg 5281 aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta 5341 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta 5401 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata 5461 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc 5521 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga 5581 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct 5641 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg 5701 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta 5761 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat 5821 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt 5881 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga 5941 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga 6001 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga 6061 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc 6121 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc 6181 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg 6241 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat 6301 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata 6361 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct 6421 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga 6481 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg 6541 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc 6601 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct 6661 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc 6721 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt 6781 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg 6841 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct 6901 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag 6961 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag 7021 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg 7081 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg 7141 gccttttgct cacatgt //