LOCUS Exported 6098 bp ds-DNA circular SYN 12-JUL-2022 DEFINITION AAV expression of Cre recombinase-activated jRCaMP1b, a red fluorescent calcium sensor protein, from the Synapsin promoter. ACCESSION . VERSION . KEYWORDS pAAV.Syn.Flex.NES-jRCaMP1b.WPRE.SV40 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6098) AUTHORS Dana H, Mohar B, Sun Y, Narayan S, Gordus A, Hasseman JP, Tsegaye G, Holt GT, Hu A, Walpita D, Patel R, Macklin JJ, Bargmann CI, Ahrens MB, Schreiter ER, Jayaraman V, Looger LL, Svoboda K, Kim DS TITLE Sensitive red protein calcium indicators for imaging neural activity. JOURNAL Elife. 2016 Mar 24;5. pii: e12727. doi: 10.7554/eLife.12727. PUBMED 27011354 REFERENCE 2 (bases 1 to 6098) AUTHORS . TITLE Direct Submission JOURNAL Exported Jul 12, 2022 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6098 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 53..182 /label=AAV2 ITR promoter 264..711 /label=hSyn promoter /note="human synapsin I promoter; confers neuron-specific expression (Kugler et al., 2003)" protein_bind complement(741..774) /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(837..870) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS complement(2120..2143) /codon_start=1 /product="Xpress(TM) epitope tag, including an enterokinase recognition and cleavage site" /label=Xpress(TM) tag /translation="DLYDDDDK" CDS complement(2147..2179) /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(2183..2200) /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" protein_bind 2283..2316 /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 2387..2420 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." misc_feature 2462..3050 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2515..2535) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(2933..2944) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(3053..3069) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3054..3070) /label=pBluescriptKS /note="For pBluescript vector" polyA_signal 3099..3220 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3110..3129) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 3164..3183 /label=SV40pA-R /note="SV40 polyA, reverse primer" rep_origin 3583..4038 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3670..3689) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3880..3901 /label=F1ori-F /note="F1 origin, forward primer" promoter 4064..4168 /gene="bla" /label=AmpR promoter CDS 4169..5029 /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(4387..4406) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5200..5788 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5689..5708 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5942..5959 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctgcgcgc 61 tcgctcgctc actgaggccg cccgggcaaa gcccgggcgt cgggcgacct ttggtcgccc 121 ggcctcagtg agcgagcgag cgcgcagaga gggagtggcc aactccatca ctaggggttc 181 cttgtagtta atgattaacc cgccatgcta cttatctacg tagccatgct ctaggaagat 241 ctctgcagag ggccctgcgt atgagtgcaa gtgggtttta ggaccaggat gaggcggggt 301 gggggtgcct acctgacgac cgaccccgac ccactggaca agcacccaac ccccattccc 361 caaattgcgc atcccctatc agagaggggg aggggaaaca ggatgcggcg aggcgcgtgc 421 gcactgccag cttcagcacc gcggacagtg ccttcgcccc cgcctggcgg cgcgcgccac 481 cgccgcctca gcactgaagg cgcgctgacg tcactcgccg gtcccccgca aactcccctt 541 cccggccacc ttggtcgcgt ccgcgccgcc gccggcccag ccggaccgca ccacgcgagg 601 cgcgagatag gggggcacgg gcgcgaccat ctgcgctgcg gcgccggcga ctcagcgctg 661 cctcagtctg cggtgggcag cggaggagtc gtgtcgtgcc tgagagcgca gtcgaattca 721 agctgctagc aaggatcccc ataacttcgt ataaagtatc ctatacgaag ttatatcaaa 781 ataggaagac caatgcttca ccatcgaccc gaattgccaa gcatcaccat cgacccataa 841 cttcgtataa tgtatgctat acgaagttat actagagctt tcacttcgct gtcatcattt 901 gtacaaactc ttcgtagttt acctgaccat ccccatcgct gtctgcttcc ctgatcattt 961 catcaacctc ttcatctgtt aacttctctc caaggtttgt catcacgtgg cgaagctctg 1021 ctgcactgat gtagccattg ccatccttat caaacacgcc gaacgcttct ctaatttctt 1081 cttcactgtc tgtgtatttc attttgcctg ccatcataat caggaactca gggaagtcga 1141 ttgtgccgtc accgtcggca tctacttcat tgatcatgtc ctgcagctct gcttctgtgg 1201 ggttctgccc cagagaccgc atcaccgtcc ccatctcctt ggttgttatt gtcccatccc 1261 cgtccttgtc aaatagggag aaagcctctt taaattctgc gatctgctct tcagtcagtt 1321 gggagtccgt aggctcccaa cctttggttt tcttttgcat tacagcacca ttgctcggga 1381 agttcacacc gcgaacctgc acgtgataca ccaggcagcc atcctccaga gacgtgtctt 1441 gcataacggt aattacgccg ccgtcttcgt aacgggtcac acgttcccag gtaaaacctt 1501 ccgggaagga ctgcttaaag aaatccggga tgcctttcgg atatttgatg aaggtacggg 1561 agccatacat aaaggacgtc gccaggatgt cgaacgcaaa cggcagcgga ccaccctcga 1621 tcactttgat acgcatagtc tgggtgccca tatacgggtt gccttcgcct tcaccagtgc 1681 atttgaactg gtggccgttt acggagcctt ccagaaccac tttcatacgc atgttctcct 1741 tgatcaggga gttcatgctc cctccggtac cgccgccacc acccaggccc acaaacttgg 1801 caactgcgtg ttcacgctgt acgacaaaca tttcgttatc gctctcctcc aggcgctcca 1861 gacggtggct gacgtaatga atggcaggca tcttaatgtt gccgacagtc tttttggagc 1921 ggtaggtggt cacgaaagaa caggacaggt gaccgccgcc atcaactttc agcgccatgt 1981 gagtgtaacc acgcagacca ccatccgctg ggtacatcat ttcggtgttg ttcgctgagc 2041 tcagccgacc tatagctctg actgcgtgac cccacttatt ccacttacgt cgcgatgagt 2101 cgaccattgt tgcgagatcc ttatcgtcat cgtcgtacag atcccgaccc atttgctgtc 2161 caccagtcat gctagccata ccatgatgat gatgatgatg agaacctcct ccagtcttgt 2221 taatatcaag tccagccaac ttaagagcaa gctcgttctg cagcatggtg gcggatccta 2281 gtataacttc gtataggata ctttatacga agttatcatt gggattcttc ctattttgat 2341 ccaagcatca ccatcgaccc tctagtccag atctcaccat cgacccataa cttcgtatag 2401 catacattat acgaagttat gtccctcgaa gaggttcgaa ttcgatatca agcttatcga 2461 taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc 2521 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg 2581 tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt 2641 gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac 2701 tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc 2761 tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct 2821 gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct 2881 cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct 2941 caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct 3001 tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgataccg 3061 tcgacctcga cccgggcggc cgcttcgagc agacatgata agatacattg atgagtttgg 3121 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat 3181 tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca 3241 ttttatgttt caggttcagg gggagatgtg ggaggttttt taaagcaagt aaaacctcta 3301 caaatgtggt aaaatcgata aggatcttcc tagagcatgg ctacgtagat aagtagcatg 3361 gcgggttaat cattaactac aaggaacccc tagtgatgga gttggccact ccctctctgc 3421 gcgctcgctc gctcactgag gccgsscgrs cwaaggtcgc ccgacgcccg ggctttgccc 3481 gggcggcctc agtgagcgag cgagcgcgca gctggcgtaa tagcgaagag gcccgcaccg 3541 atcgcccttc ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg 3601 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc 3661 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc 3721 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg 3781 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg 3841 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg 3901 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt 3961 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa 4021 tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg 4081 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat 4141 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat 4201 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt 4261 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag 4321 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa 4381 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg 4441 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct 4501 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac 4561 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca 4621 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat 4681 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact 4741 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc 4801 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga 4861 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg 4921 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg 4981 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca 5041 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta 5101 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca 5161 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg 5221 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga 5281 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa 5341 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc 5401 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg 5461 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac 5521 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct 5581 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc 5641 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg 5701 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg 5761 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct 5821 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga 5881 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg 5941 cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcamwccry cwctaggggt 6001 tccttgtagt taatgattaa cccgccatgc tacttatcta cgtagccatg ctctaggaag 6061 atccttatcg attttaccac atttgtagag gttttact //