LOCUS Exported 5933 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION AAV-mediated expression of ultrasensitive protein calcium sensor under the Syn promoter, Cre-dependent expression. ACCESSION . VERSION . KEYWORDS pGP-AAV-syn-FLEX-jGCaMP7c variant 1513-WPRE SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5933) AUTHORS Dana H, Sun Y, Mohar B, Hulse BK, Kerlin AM, Hasseman JP, Tsegaye G, Tsang A, Wong A, Patel R, Macklin JJ, Chen Y, Konnerth A, Jayaraman V, Looger LL, Schreiter ER, Svoboda K, Kim DS TITLE High-performance calcium sensors for imaging activity in neuronal populations and microcompartments. JOURNAL Nat Methods. 2019 Jul;16(7):649-657. doi: 10.1038/s41592-019-0435-6. Epub 2019 Jun 17. PUBMED 31209382 REFERENCE 2 (bases 1 to 5933) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5933 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 45..174 /label=AAV2 ITR promoter 256..703 /label=hSyn promoter /note="human synapsin I promoter; confers neuron-specific expression (Kugler et al., 2003)" protein_bind complement(733..766) /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(829..862) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind 1432..1451 /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 1691..1712 /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1818..1839) /label=EGFP-C /note="EGFP, forward primer" CDS complement(2052..2108) /codon_start=1 /product="calmodulin-binding peptide from avian smooth muscle myosin light chain kinase" /label=calmodulin-binding peptide /translation="RRKWNKTGHAVRAIGRLSS" regulatory 2120..2129 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS complement(2136..2159) /codon_start=1 /product="Xpress(TM) epitope tag, including an enterokinase recognition and cleavage site" /label=Xpress(TM) tag /translation="DLYDDDDK" CDS complement(2163..2195) /codon_start=1 /product="leader peptide from bacteriophage T7 gene 10" /label=T7 tag (gene 10 leader) /note="promotes efficient translation in E. coli" /translation="MASMTGGQQMG" CDS complement(2199..2216) /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" regulatory 2222..2231 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" protein_bind 2242..2275 /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 2346..2379 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." misc_feature 2421..3009 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2474..2494) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(2892..2903) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3052..3173 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3063..3082) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 3117..3136 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 3335..3464 /label=AAV2 ITR primer_bind complement(3530..3547) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(3701..4289) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3781..3800) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(4460..5320) /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 5083..5102 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5321..5425) /gene="bla" /label=AmpR promoter rep_origin complement(5451..5906) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5588..5609) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 5800..5819 /label=F1ori-R /note="F1 origin, reverse primer" ORIGIN 1 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctgcgc gctcgctcgc 61 tcactgaggc cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag 121 tgagcgagcg agcgcgcaga gagggagtgg ccaactccat cactaggggt tccttgtagt 181 taatgattaa cccgccatgc tacttatcta cgtagccatg ctctaggaag atctctgcag 241 agggccctgc gtatgagtgc aagtgggttt taggaccagg atgaggcggg gtgggggtgc 301 ctacctgacg accgaccccg acccactgga caagcaccca acccccattc cccaaattgc 361 gcatccccta tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt gcgcactgcc 421 agcttcagca ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc accgccgcct 481 cagcactgaa ggcgcgctga cgtcactcgc cggtcccccg caaactcccc ttcccggcca 541 ccttggtcgc gtccgcgccg ccgccggccc agccggaccg caccacgcga ggcgcgagat 601 aggggggcac gggcgcgacc atctgcgctg cggcgccggc gactcagcgc tgcctcagtc 661 tgcggtgggc agcggaggag tcgtgtcgtg cctgagagcg cagtcgaatt caagctgcta 721 gcaaggatcc ccataacttc gtataaagta tcctatacga agttatatca aaataggaag 781 accaatgctt caccatcgac ccgaattgcc aagcatcacc atcgacccat aacttcgtat 841 aatgtatgct atacgaagtt atactagagc tttcacttcg ctgtcatcat ttgtacaaac 901 tcttcgtagt ttacctgacc atccccatcg atgtctgctt ccctgatcat ttcatcaacc 961 tcttcatctg ttaacttctc tccaaggttg ttcatcacgt ggcgaagctc tgctgcactg 1021 atgtagccat tgccatcctt atcaaacaca ccgaacgctt ctctaatttc ttcttcactg 1081 tccctgtagc tcccttttct tgccatcatt gtcaggaact cagggaagtc gattgtgccg 1141 tcaccgtcgg catctacttc attgatcatg tcctgcagct ctgcttctgt ggggttctgc 1201 cccagagaac gcatcaccgt ccccagctcc ttggttgtta ttgtcccatc cccgtccttg 1261 tcaaataggg agaaagcctc tttaaattct gcgatctgct cttcagtcag ttggtcagga 1321 aggttgtact ccagcttgtg ccccaggatg ttgccgtcct ccttgaagtc gatgcccttc 1381 agctcgatgc ggttcaccag ggtgtcgccc tcgaacttca cctcggcgcg ggtcttgtag 1441 ttgccgtcgt ccttgaagaa gatggtgcgc tcctggatgt agccttcggg catggcggac 1501 ttgaagaagt cgtgctgctt catgtggtcg gggtagcggc tgaagcactg cacgccgtag 1561 gtcagggtgg tcacgagggt gggccagggc acgggcagct tgccggtggt gcagatgaac 1621 ttcagggtca gcttgccgta ggtggcatcg ccctcaccct cgccggacac gctgaacttg 1681 tggccgttta cgtcgccgtc cagctcgacc aggatgggca ccaccccggt gaacagctcc 1741 tcgcccttgc tcaccatgct ccctccggta ccgcccttgt acagctcgtc catgccgaga 1801 gtgatcccgg cggcggtcac gaactccagc aggaccatgt gatcgcgctt ctcgttgggg 1861 tctttcgaaa gtttggactg cacgctcagg tagtggttgt cgggcagcag cacggggccg 1921 tcgccgatgg gggtgttctg ctggtagtgg taggcgagct gcacgccgcc gtcctcgatg 1981 ttgtggcgga tcttgaagtt cgccttgatg ccgttcttct gcttgtcggc cttgatatag 2041 acgttaggct gtgagctcag ccgacctata gctctgactg cgtgacctgt cttattccac 2101 ttacgacgtg atgagtcgac catggtggcg agatccttat cgtcatcgtc gtacagatcc 2161 cgacccattt gctgtccacc agtcatgcta gccataccat gatgatgatg atgatgagaa 2221 cccatggtgg cggatcctag tataacttcg tataggatac tttatacgaa gttatcattg 2281 ggattcttcc tattttgatc caagcatcac catcgaccct ctagtccaga tctcaccatc 2341 gacccataac ttcgtatagc atacattata cgaagttatg tccctcgaag aggttcgaat 2401 tcgatatcaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg 2461 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt 2521 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc 2581 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt 2641 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga 2701 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct 2761 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat 2821 cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct 2881 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc 2941 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg 3001 cctccccgca tcgataccgt cgacccgggc ggccgcttcg agcagacatg ataagataca 3061 ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa 3121 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca 3181 acaattgcat tcattttatg tttcaggttc agggggagat gtgggaggtt ttttaaagca 3241 agtaaaacct ctacaaatgt ggtaaaatcg ataaggatct tcctagagca tggctacgta 3301 gataagtagc atggcgggtt aatcattaac tacaaggaac ccctagtgat ggagttggcc 3361 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc 3421 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcat taatgaatcg 3481 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg 3541 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa 3601 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc 3661 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc 3721 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat 3781 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc 3841 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct 3901 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg 3961 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc 4021 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga 4081 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa 4141 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta 4201 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc 4261 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg 4321 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga 4381 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg 4441 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct 4501 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg 4561 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc 4621 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa 4681 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc 4741 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt 4801 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc 4861 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt 4921 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc 4981 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt 5041 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata 5101 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga 5161 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag 5221 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa 5281 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt 5341 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga 5401 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct aaattgtaag 5461 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca 5521 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag 5581 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg 5641 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt 5701 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag 5761 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc 5821 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc 5881 gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca act //