LOCUS Exported 5932 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION AAV expression of ultrasensitive protein calcium sensor from Synapsin promoter. ACCESSION . VERSION . KEYWORDS pAAV.Syn.GCaMP6s.WPRE.SV40 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5932) AUTHORS Chen TW, Wardill TJ, Sun Y, Pulver SR, Renninger SL, Baohan A, Schreiter ER, Kerr RA, Orger MB, Jayaraman V, Looger LL, Svoboda K, Kim DS TITLE Ultrasensitive fluorescent proteins for imaging neuronal activity. JOURNAL Nature. 2013 Jul 18;499(7458):295-300. doi: 10.1038/nature12354. PUBMED 23868258 REFERENCE 2 (bases 1 to 5932) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5932 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 211..658 /label=hSyn promoter /note="human synapsin I promoter; confers neuron-specific expression (Kugler et al., 2003)" regulatory 690..699 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 696..2048 /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 792..801 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" primer_bind 1082..1103 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(1209..1230) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1470..1489) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" misc_feature 2061..2649 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2114..2134) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(2532..2543) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(2652..2668) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2653..2669) /label=pBluescriptKS /note="For pBluescript vector" polyA_signal 2698..2819 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2709..2728) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2763..2782 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 2981..3110 /label=AAV2 ITR primer_bind complement(3129..3146) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(3129..3145) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3138..3160) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 3287..3742 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3374..3393) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3584..3605 /label=F1ori-F /note="F1 origin, forward primer" promoter 3768..3872 /gene="bla" /label=AmpR promoter CDS 3873..4733 /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(4091..4110) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 4904..5492 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5393..5412 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5646..5663 /label=L4440 /note="L4440 vector, forward primer" protein_bind 5780..5801 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5816..5846 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5854..5870 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." primer_bind 5859..5881 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 5878..5894 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 5878..5894 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" repeat_region 5932..129 /label=AAV2 ITR ORIGIN 1 tgcgcgctcg ctcgctcact gaggccgccc gggcaaagcc cgggcgtcgg gcgacctttg 61 gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg agtggccaac tccatcacta 121 ggggttcctt gtagttaatg attaacccgc catgctactt atctacgtag ccatgctcta 181 ggaagatctc tgcagagggc cctgcgtatg agtgcaagtg ggttttagga ccaggatgag 241 gcggggtggg ggtgcctacc tgacgaccga ccccgaccca ctggacaagc acccaacccc 301 cattccccaa attgcgcatc ccctatcaga gagggggagg ggaaacagga tgcggcgagg 361 cgcgtgcgca ctgccagctt cagcaccgcg gacagtgcct tcgcccccgc ctggcggcgc 421 gcgccaccgc cgcctcagca ctgaaggcgc gctgacgtca ctcgccggtc ccccgcaaac 481 tccccttccc ggccaccttg gtcgcgtccg cgccgccgcc ggcccagccg gaccgcacca 541 cgcgaggcgc gagatagggg ggcacgggcg cgaccatctg cgctgcggcg ccggcgactc 601 agcgctgcct cagtctgcgg tgggcagcgg aggagtcgtg tcgtgcctga gagcgcagtc 661 gaattcaagc tgctagcaag gatccacccg ccaccatggg ttctcatcat catcatcatc 721 atggtatggc tagcatgact ggtggacagc aaatgggtcg ggatctgtac gacgatgacg 781 ataaggatct cgccaccatg gtcgactcat cacgtcgtaa gtggaataag acaggtcacg 841 cagtcagagc tataggtcgg ctgagctcac tcgagaacgt ctatatcaag gccgacaagc 901 agaagaacgg catcaaggcg aacttccaca tccgccacaa catcgaggac ggcggcgtgc 961 agctcgccta ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg 1021 acaaccacta cctgagcgtg cagtccaaac tttcgaaaga ccccaacgag aagcgcgatc 1081 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt 1141 acaagggcgg taccggaggg agcatggtga gcaagggcga ggagctgttc accggggtgg 1201 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg 1261 agggtgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca 1321 agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca 1381 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct 1441 acatccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg 1501 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg 1561 aggacggcaa catcctgggg cacaagctgg agtacaacct gccggaccaa ctgactgaag 1621 agcagatcgc agaatttaaa gaggctttct ccctatttga caaggacggg gatgggacaa 1681 taacaaccaa ggagctgggg acggtgatgc ggtctctggg gcagaacccc acagaagcag 1741 agctgcagga catgatcaat gaagtagatg ccgacggtga cggcacaatc gacttccctg 1801 agttcctgac aatgatggca agaaaaatga aatacaggga cacggaagaa gaaattagag 1861 aagcgttcgg tgtgtttgat aaggatggca atggctacat cagtgcagca gagcttcgcc 1921 acgtgatgac aaaccttgga gagaagttaa cagatgaaga ggttgatgaa atgatcaggg 1981 aagcagacat cgatggggat ggtcaggtaa actacgaaga gtttgtacaa atgatgacag 2041 cgaagtgaaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg 2101 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt 2161 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc 2221 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt 2281 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga 2341 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct 2401 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat 2461 cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct 2521 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc 2581 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg 2641 cctccccgca tcgataccgt cgacctcgac ccgggcggcc gcttcgagca gacatgataa 2701 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt 2761 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta 2821 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggagatgtgg gaggtttttt 2881 aaagcaagta aaacctctac aaatgtggta aaatcgataa ggatcttcct agagcatggc 2941 tacgtagata agtagcatgg cgggttaatc attaactaca aggaacccct agtgatggag 3001 ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc 3061 cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ccttaattaa 3121 cctaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa 3181 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc 3241 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc 3301 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc 3361 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt 3421 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac 3481 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag 3541 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa 3601 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg 3661 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac 3721 aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta 3781 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat 3841 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc 3901 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga 3961 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca 4021 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt 4081 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg 4141 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc 4201 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata 4261 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt 4321 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag 4381 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca 4441 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg 4501 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg 4561 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag 4621 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg 4681 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag 4741 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga 4801 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt 4861 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc 4921 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc 4981 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac 5041 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac 5101 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt 5161 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct 5221 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat 5281 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt 5341 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg 5401 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt 5461 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt 5521 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg 5581 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg 5641 agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc 5701 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg 5761 gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac 5821 actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag 5881 gaaacagcta tgaccatgat tacgccagat ttaattaagg ccttaattag gc //