LOCUS Exported 6485 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION AAV expression of ultrasensitive protein calcium sensor from the CamKII promoter. ACCESSION . VERSION . KEYWORDS AAV.CamKII.GCaMP6s.WPRE.SV40 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6485) TITLE AAV plasmids JOURNAL Unpublished REFERENCE 2 (bases 1 to 6485) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6485 /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 160..1448 /label=CaMKII promoter /note="promoter for the mouse calcium/calmodulin-dependent kinase II alpha subunit" regulatory 1473..1482 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1479..2831 /codon_start=1 /product="improved fluorescent protein-based calcium sensor (Chen et al., 2013)" /label=GCaMP6s /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK TGHAVRAIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGP VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRS LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK" regulatory 1575..1584 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" primer_bind 1865..1886 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(1992..2013) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2253..2272) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" misc_feature 2844..3432 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2897..2917) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3315..3326) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3475..3596 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3486..3505) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 3540..3559 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 3748..3888 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 3748..3877 /label=AAV2 ITR rep_origin 3963..4418 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4050..4069) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4260..4281 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4435..4454) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4554..4576 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4614..4632) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4700..4804 /gene="bla" /label=AmpR promoter CDS 4805..5665 /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(5023..5042) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5836..6424 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6325..6344 /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 cgcgtttaac attatggcct taggtcactt 181 catctccatg gggttcttct tctgattttc tagaaaatga gatgggggtg cagagagctt 241 cctcagtgac ctgcccaggg tcacatcaga aatgtcagag ctagaacttg aactcagatt 301 actaatctta aattccatgc cttgggggca tgcaagtacg atatacagaa ggagtgaact 361 cattagggca gatgaccaat gagtttagga aagaagagtc cagggcaggg tacatctaca 421 ccacccgccc agccctgggt gagtccagcc acgttcacct cattatagtt gcctctctcc 481 agtcctacct tgacgggaag cacaagcaga aactgggaca ggagccccag gagaccaaat 541 cttcatggtc cctctgggag gatgggtggg gagagctgtg gcagaggcct caggaggggc 601 cctgctgctc agtggtgaca gataggggtg agaaagcaga cagagtcatt ccgtcagcat 661 tctgggtctg tttggtactt cttctcacgc taaggtggcg gtgtgatatg cacaatggct 721 aaaaagcagg gagagctgga aagaaacaag gacagagaca gaggccaagt caaccagacc 781 aattcccaga ggaagcaaag aaaccattac agagactaca agggggaagg gaaggagaga 841 tgaattagct tcccctgtaa accttagaac ccagctgttg ccagggcaac ggggcaatac 901 ctgtctcttc agaggagatg aagttgccag ggtaactaca tcctgtcttt ctcaaggacc 961 atcccagaat gtggcaccca ctagccgtta ccatagcaac tgcctctttg ccccacttaa 1021 tcccatcccg tctgttaaaa gggccctata gttggaggtg ggggaggtag gaagagcgat 1081 gatcacttgt ggactaagtt tgttcgcatc cccttctcca accccctcag tacatcaccc 1141 tgggggaaca gggtccactt gctcctgggc ccacacagtc ctgcagtatt gtgtatataa 1201 ggccagggca aagaggagca ggttttaaag tgaaaggcag gcaggtgttg gggaggcagt 1261 taccggggca acgggaacag ggcgtttcgg aggtggttgc catggggacc tggatgctga 1321 cgaaggctcg cgaggctgtg agcagccaca gtgccctgct cagaagcccc aagctcgtca 1381 gtcaagccgg ttctccgttt gcactcagga gcacgggcag gcgagtggcc cctagttctg 1441 ggggcagctc tagagcggta ccggatccac ccgccaccat gggttctcat catcatcatc 1501 atcatggtat ggctagcatg actggtggac agcaaatggg tcgggatctg tacgacgatg 1561 acgataagga tctcgccacc atggtcgact catcacgtcg taagtggaat aagacaggtc 1621 acgcagtcag agctataggt cggctgagct cactcgagaa cgtctatatc aaggccgaca 1681 agcagaagaa cggcatcaag gcgaacttcc acatccgcca caacatcgag gacggcggcg 1741 tgcagctcgc ctaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc 1801 ccgacaacca ctacctgagc gtgcagtcca aactttcgaa agaccccaac gagaagcgcg 1861 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc 1921 tgtacaaggg cggtaccgga gggagcatgg tgagcaaggg cgaggagctg ttcaccgggg 1981 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg 2041 gcgagggtga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg 2101 gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct 2161 tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag 2221 gctacatcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg 2281 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca 2341 aggaggacgg caacatcctg gggcacaagc tggagtacaa cctgccggac caactgactg 2401 aagagcagat cgcagaattt aaagaggctt tctccctatt tgacaaggac ggggatggga 2461 caataacaac caaggagctg gggacggtga tgcggtctct ggggcagaac cccacagaag 2521 cagagctgca ggacatgatc aatgaagtag atgccgacgg tgacggcaca atcgacttcc 2581 ctgagttcct gacaatgatg gcaagaaaaa tgaaatacag ggacacggaa gaagaaatta 2641 gagaagcgtt cggtgtgttt gataaggatg gcaatggcta catcagtgca gcagagcttc 2701 gccacgtgat gacaaacctt ggagagaagt taacagatga agaggttgat gaaatgatca 2761 gggaagcaga catcgatggg gatggtcagg taaactacga agagtttgta caaatgatga 2821 cagcgaagtg aaagcttatc gataatcaac ctctggatta caaaatttgt gaaagattga 2881 ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt 2941 tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt 3001 tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg 3061 tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg 3121 ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc 3181 gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaaat 3241 catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct 3301 tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg 3361 ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg 3421 ccgcctcccc gcatcgatac cgtcgacccg ggcggccgct tcgagcagac atgataagat 3481 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg 3541 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca 3601 acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag gttttttaaa 3661 gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga tcttcctaga gcatggctac 3721 gtaaccacgt gcggaccgag cggccgcagg aacccctagt gatggagttg gccactccct 3781 ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct 3841 ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc 3901 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata 3961 gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac 4021 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc 4081 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt 4141 tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg 4201 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag 4261 tggactcttg ttccaaactg gaacaacact caaccctatc tcgggctatt cttttgattt 4321 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt 4381 taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat 4441 ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc 4501 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag 4561 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt 4621 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg 4681 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa 4741 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa 4801 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct 4861 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg 4921 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg 4981 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt 5041 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga 5101 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga 5161 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac 5221 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg 5281 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac 5341 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct 5401 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct 5461 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg 5521 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat 5581 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg 5641 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat 5701 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct 5761 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa 5821 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa 5881 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc 5941 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta 6001 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct 6061 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg 6121 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag 6181 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc 6241 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg 6301 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt 6361 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg 6421 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca 6481 catgt //