LOCUS Exported 7388 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION AAV vector for Cre-dependent expression of Venus-tagged parapinopsin driven by the CAG promoter. Used for photoswitchable control of inhibitory GPCR signaling cascades.. ACCESSION . VERSION . KEYWORDS pAAV-CAG-DIO-PPO-Venus SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7388) AUTHORS Copits BA, O'Neill PR, Gowrishankar R, Yoo JJ, Meshik X, Parker KE, Spangler SM, Vasquez AM, Elerding AJ, Stander MC, Kalyanaraman V, Vogt SK, Samineni VK, Gautam N, Sunahara RK, Gereau RW, Bruchas MR. TITLE A photoswitchable GPCR-based opsin for presynaptic silencing. JOURNAL BioRxiv REFERENCE 2 (bases 1 to 7388) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7388 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 432..454 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 516..535 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" protein_bind 586..619 /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 670..703 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS complement(737..1456) /codon_start=1 /product="Venus YFP with monomerizing A206K mutation (Nagai et al., 2002; Kremers et al., 2006)" /label=mVenus /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(782..803) /label=EGFP-C /note="EGFP, forward primer" primer_bind 1131..1150 /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 1390..1411 /label=EGFP-N /note="EGFP, reverse primer" regulatory 1453..1462 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" protein_bind complement(2514..2547) /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(2598..2631) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." misc_feature 2656..3244 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2709..2729) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3127..3138) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(3271..3288) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 3277..3501 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" promoter complement(3711..4040) /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3725..3860 /label=SV40 ori /note="SV40 origin of replication" primer_bind complement(3779..3798) /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind 4025..4045 /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" primer_bind complement(4115..4132) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4286..4874) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4366..4385) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5045..5905) /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 5668..5687 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5906..6010) /gene="bla" /label=AmpR promoter primer_bind 6078..6096 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(6134..6156) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 6256..6275 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 6489..6508 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" promoter 6798..6816 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind 6798..6815 /label=SP6 /note="SP6 promoter, forward primer" repeat_region 6837..6966 /label=AAV2 ITR ORIGIN 1 gcgccaggcg gggcggggcg gggcgagggg cggggcgggg cgaggcggag aggtgcggcg 61 gcagccaatc agagcggcgc gctccgaaag tttcctttta tggcgaggcg gcggcggcgg 121 cggccctata aaaagcgaag cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc 181 gtgccccgct ccgccgccgc ctcgcgccgc ccgccccggc tctgactgac cgcgttactc 241 ccacaggtga gcgggcggga cggcccttct cctccgggct gtaattagcg cttggtttaa 301 tgacggcttg tttcttttct gtggctgcgt gaaagccttg aggggctccg ggagggccct 361 ttgtgcgggg ggagcggctc ggggctgtcc gcggggggac ggctgccttc gggggggacg 421 gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct ctgctaacca 481 tgttcatgcc ttcttctttt tcctacagct cctgggcaac gtgctggtta ttgtgctgtc 541 tcatcatttt ggcaaagaat tggatcctct agagtcgact ccggaataac ttcgtatagg 601 atactttata cgaagttatg cagaatggta gctggattgt agctgctatt agcaatatga 661 aacctcttaa taacttcgta tagcatacat tatacgaagt tatggcgcgc cagcgggttt 721 aaacgggccc tctagattac ttgtacagct cgtccatgcc gagagtgatc ccggcggcgg 781 tcacgaactc cagcaggacc atgtgatcgc gcttctcgtt ggggtctttg ctcagtttgg 841 actggtagct caggtagtgg ttgtcgggca gcagcacggg gccgtcgccg atgggggtgt 901 tctgctggta gtggtcggcg agctgcacgc cgccgtcctc gatgttgtgg cggatcttga 961 agttggcctt gatgccgttc ttctgcttgt cggcggtgat atagacgttg tggctgttgt 1021 agttgtactc cagcttgtgc cccaggatgt tgccgtcctc cttgaagtcg atgcccttca 1081 gctcgatgcg gttcaccagg gtgtcgccct cgaacttcac ctcggcgcgg gtcttgtagt 1141 tgccgtcgtc cttgaagaag atggtgcgct cctggacgta gccttcgggc atggcggact 1201 tgaagaagtc gtgctgcttc atgtggtcgg ggtagcgggc gaagcactgc aggccgtagc 1261 ccagggtggt cacgagggtg ggccagggca cgggcagctt gccggtggtg cagatcagct 1321 tcagggtcag cttgccgtag gtggcatcgc cctcgccctc gccggacacg ctgaacttgt 1381 ggccgtttac gtcgccgtcc agctcgacca ggatgggcac caccccggtg aacagctcct 1441 cgcccttgct caccatggcg gccgcgctcg gggagacctg ccccggcgcg gacgccaggg 1501 tcacgctggt ggccgacgtg gaggccgccg tcgccgactc cgaggaaggc tctgcccagg 1561 ggttgcgtcc gcagagcagg aagggcacgg cgcagtcccg gaactggcgg ttcatgaaga 1621 tgtagatgat ggggttgtag accgtgctcg tcttggtcag gtacatgggc agtgtggcga 1681 tgaccgggtc gatgtacacg tcgggcttgg tcaccacgat catggcgaag agcgcgtagg 1741 gcagccagca gacgaggaag gccaccacca tgaccaccac catgcgcgac acctgcgcct 1801 ccgccttggc ggtgctgccg ctctcgccca tccccagctt ggccacctgg tggagggtcc 1861 acatgaggcg gccgtacgcc accacgatga cgaggaaggg gatggcgaag cagaaggcga 1921 agtagctcgt aatgtacgac acgttggcgg ggtcgcggct gtaccagtcg ggcgcgcacg 1981 acgtccgcac gccctccagc tcgtagctgc cccagccgaa gagcggcggc gtgttccaca 2041 cgaacgacca gagccaggcc caggcgatgc ccagcagcgc gtggcgccgc gtgaacatca 2101 gcgtgcccag cggcttgcac accaccacga agcgatcgac ggcgatcacg gcgatcgtgc 2161 acagagcggc gatgccgaag aaagcgacgg cgaacccttc gatgacgcag ccgacgcggc 2221 cgaggttgaa gtacccgacg gcgttggtct ccacgacgag gctggcgccg aacaccgtga 2281 cgcccaggtc cgccacggcg aggttgacga gcgagaagtt gagcgggtgg cgtagctggc 2341 ggtggcgcag ggtgacgatg acgacggtgc tgttgagcac gagcgaggcg atggtgaaca 2401 cggccatgat cacggcgagg atggtgaagc cgtagcgggg catcaacggg acctcgccgt 2461 tgggcaggag gtcgagcgag gtcaagttct ccatgaattc caccggtgct agcataactt 2521 cgtataaagt atcctatacg aagttatttg ccttaaccca gaaattatca ctgttattct 2581 ttagaatggt gcaaagaata acttcgtata atgtatgcta tacgaagtta tgaattcgat 2641 atcaagctta tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt 2701 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat 2761 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct 2821 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct 2881 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc 2941 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg 3001 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc 3061 tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac 3121 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg 3181 cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc 3241 ccgcatcgat accgtcgact cgctgatcag cctcgactgt gccttctagt tgccagccat 3301 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc 3361 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg 3421 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg 3481 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc tcgactagag 3541 catggctacg tagataagta gcatggcggg ttaatcatta actacaagga acccctagtg 3601 atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag 3661 gtcgcccgac gcccgggcgg cctcagtgag cgagcgagcg cgcagagctt tttgcaaaag 3721 cctaggcctc caaaaaagcc tcctcactac ttctggaata gctcagaggc cgaggcggcc 3781 tcggcctctg cataaataaa aaaaattagt cagccatggg gcggagaatg ggcggaactg 3841 ggcggagtta ggggcgggat gggcggagtt aggggcggga ctatggttgc tgactaattg 3901 agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc acacctggtt 3961 gctgactaat tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt 4021 ccacacccta actgacacac attccacagc tgcattaatg aatcggccaa cgcgcgggga 4081 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg 4141 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag 4201 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc 4261 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca 4321 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt 4381 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc 4441 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc 4501 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc 4561 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact 4621 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg 4681 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta 4741 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca 4801 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa 4861 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg 4921 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc 4981 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg 5041 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat 5101 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg 5161 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa 5221 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca 5281 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc 5341 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt 5401 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa 5461 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat 5521 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct 5581 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga 5641 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag 5701 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga 5761 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca 5821 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg 5881 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc 5941 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag 6001 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca 6061 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg 6121 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag 6181 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg 6241 gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat tcgacgctct 6301 cccttatgcg actcctgcat taggaagcag cccagtagta ggttgaggcc gttgagcacc 6361 gccgccgcaa ggaatggtgc atgcaaggag atggcgccca acagtccccc ggccacgggg 6421 cctgccacca tacccacgcc gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct 6481 tccccatcgg tgatgtcggc gatataggcg ccagcaaccg cacctgtggc gccggtgatg 6541 ccggccacga tgcgtccggc gtagaggatc tggctagcga tgaccctgct gattggttcg 6601 ctgaccattt ccgggtgcgg gacggcgtta ccagaaactc agaaggttcg tccaaccaaa 6661 ccgactctga cggcagttta cgagagagat gatagggtct gcttcagtaa gccagatgct 6721 acacaattag gcttgtacat attgtcgtta gaacgcggct acaattaata cataacctta 6781 tgtatcatac acatacgatt taggtgacac tatagaatac acggaattaa ttctagctgc 6841 gcgctcgctc gctcactgag gccgcccggg caaagcccgg gcgtcgggcg acctttggtc 6901 gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc atcactaggg 6961 gttccttgta gttaatgatt aacccgccat gctacttatc tacgtagcca tgctctaggt 7021 accattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg 7081 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca 7141 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc 7201 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc 7261 tattaccatg gtcgaggtga gccccacgtt ctgcttcact ctccccatct cccccccctc 7321 cccaccccca attttgtatt tatttatttt ttaattattt tgtgcagcga tgggggcggg 7381 gggggggg //