LOCUS Exported 6412 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION AAV expression of Cre recombinase-activated jRCaMP1a, a red fluorescent calcium sensor protein, from CAG promoter. ACCESSION . VERSION . KEYWORDS pAAV.CAG.Flex.NES-jRCaMP1a.WPRE.SV40 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6412) 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 6412) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6412 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..130 /label=AAV2 ITR promoter 441..718 /label=chicken beta-actin promoter primer_bind 997..1019 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 1081..1100 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" protein_bind complement(1135..1168) /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(1231..1264) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." CDS complement(2430..2486) /codon_start=1 /product="calmodulin-binding peptide from avian smooth muscle myosin light chain kinase" /label=calmodulin-binding peptide /translation="RRKWNKTGHAVRAIGRLSS" CDS complement(2514..2537) /codon_start=1 /product="Xpress(TM) epitope tag, including an enterokinase recognition and cleavage site" /label=Xpress(TM) tag /translation="DLYDDDDK" CDS complement(2541..2573) /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(2577..2594) /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" protein_bind 2677..2710 /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 2781..2814 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." misc_feature 2856..3444 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(2909..2929) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3327..3338) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3487..3608 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3498..3517) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 3552..3571 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 3770..3899 /label=AAV2 ITR primer_bind complement(3965..3982) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4136..4724) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4216..4235) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(4895..5755) /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 5518..5537 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5756..5860) /gene="bla" /label=AmpR promoter rep_origin complement(5886..6341) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(6023..6044) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 6235..6254 /label=F1ori-R /note="F1 origin, reverse primer" ORIGIN 1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt 61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact 121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct 181 aggaagagta ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac 241 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca 301 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg 361 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt 421 agtcatcgct attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc 481 ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat 541 gggggcgggg gggggggggg ggcgcgcgcc aggcggggcg gggcggggcg aggggcgggg 601 cggggcgagg cggagaggtg cggcggcagc caatcagagc ggcgcgctcc gaaagtttcc 661 ttttatggcg aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc ggcgggcggg 721 agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc 781 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc 841 gggctgtaat tagcgcttgg tttaatgacg gcttgtttct tttctgtggc tgcgtgaaag 901 ccttgagggg ctccgggagg gccctttgtg cggggggagc ggctcggggc tgtccgcggg 961 gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg 1021 gcggctctag agcctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg 1081 gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattggat ccccataact 1141 tcgtataaag tatcctatac gaagttatat caaaatagga agaccaatgc ttcaccatcg 1201 acccgaattg ccaagcatca ccatcgaccc ataacttcgt ataatgtatg ctatacgaag 1261 ttatactaga gctttcactt cgctgtcatc atttgtacaa actcttcgta gtttacctga 1321 ccatccccat cgctgtctgc ttccctgatc atttcatcaa cctcttcatc tgttaacttc 1381 tctccaaggt ttgtcatcac gtggcgaagc tctgctgcac tgatgtagcc attgccatcc 1441 ttatcaaaca cgccgaacgc ttctctaatt tcttcttcac tgtctgtgta tttcattttt 1501 cttgccatca taatcaggaa ctcagggaag tcgattgtgc cgtcaccgtc ggcatctact 1561 tcattgatca tgtcctgcag ctctgcttct gtggggttct gccccagaga ccgcatcacc 1621 gtccccagct ccttggttgt tattgtccca tccccgtcct tgtcaaatag ggagaaagcc 1681 tctttaaatt ctgcgatctg ctcttcagtg gtttgggagt ccgtaggctc ccaacctttg 1741 gttttctttt gcattacagc accattgctc gggaagttca caccgcgaac ctgcacgtga 1801 tacaccaggc agccatcctc cagagacgtg tcttgcataa cggtaattac gccgccgtct 1861 tcgtaacggg tcacacgttc ccaggtaaaa ccttccggga aggactgctt aaagaaatcc 1921 gggatgcctt tcggatattt gatgaaggta cgggagccat acataaagga cgtcgccagg 1981 atgtcgaacg caaacggcag cggaccaccc tcgatcactt tgatacgcat agtctgggtg 2041 cccatatacg ggttgccttc gccttcacca gtgcatttga actggtggcc gtttacggag 2101 ccttccagaa ccactttcat acgcatgttc tccttgatca gggagttcat gctccctccg 2161 gtaccgccgc caccacccag gcccacaaac ttggcaactg cgtgttcacg ctgtacgaca 2221 aacatttcgt tatcgctctc ctccaggcgc tccagacggt ggctgacgct atgaatggca 2281 ggcatcttaa tgttgccgac agtctttttg gagcggtagg tggtcacgaa agaacaggac 2341 aggtgaccgc cgccatcaac tttcagcgcc atgtgagtgt aaccacgcag accaccatcc 2401 gctgggtaca tcatttcgca gttgatcgct gagctcagcc gacctatagc tctgactgcg 2461 tgacctgtct tattccactt acgacgtgat gagtcgacca ttgttgcgag atccttatcg 2521 tcatcgtcgt acagatcccg acccatttgc tgtccaccag tcatgctagc cataccatga 2581 tgatgatgat gatgagaacc tcctccagtc ttgttaatat caagtccagc caacttaaga 2641 gcaagctcgt tctgcagcat ggtggcggat cctagtataa cttcgtatag gatactttat 2701 acgaagttat cattgggatt cttcctattt tgatccaagc atcaccatcg accctctagt 2761 ccagatctca ccatcgaccc ataacttcgt atagcataca ttatacgaag ttatgtccct 2821 cgaagaggtt cgaattcgat atcaagctta tcgataatca acctctggat tacaaaattt 2881 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg 2941 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt 3001 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg 3061 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc 3121 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg 3181 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt 3241 tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc 3301 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg 3361 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga 3421 tctccctttg ggccgcctcc ccgcatcgat accgtcgacc cgggcggccg cttcgagcag 3481 acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat 3541 gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata 3601 aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg gagatgtggg 3661 aggtttttta aagcaagtaa aacctctaca aatgtggtaa aatcgataag gatcttccta 3721 gagcatggct acgtagataa gtagcatggc gggttaatca ttaactacaa ggaaccccta 3781 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca 3841 aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc 3901 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg 3961 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc 4021 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt 4081 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc 4141 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa 4201 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc 4261 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg 4321 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc 4381 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc 4441 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca 4501 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact 4561 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg 4621 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt 4681 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct 4741 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga 4801 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa 4861 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac 4921 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga 4981 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc 5041 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca 5101 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta 5161 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg 5221 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc 5281 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg 5341 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt 5401 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt 5461 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata 5521 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc 5581 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac 5641 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa 5701 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct 5761 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat 5821 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc 5881 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag 5941 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac 6001 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga 6061 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc 6121 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg 6181 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa 6241 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac 6301 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct 6361 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc ag //