LOCUS Exported 5308 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression of low affinity calcium-sensing fluorescent protein. ACCESSION . VERSION . KEYWORDS pGP-CMV-GCaMP6-150 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5308) AUTHORS de Juan-Sanz J, Holt GT, Schreiter ER, de Juan F, Kim DS, Ryan TA TITLE Axonal Endoplasmic Reticulum Ca2+ Content Controls Release Probability in CNS Nerve Terminals. JOURNAL Neuron. 2017 Jan 30. pii: S0896-6273(17)30034-X. doi: 10.1016/j.neuron.2017.01.010. PUBMED 28162809 REFERENCE 2 (bases 1 to 5308) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5308 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 61..364 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 365..568 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 519..539 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 616..625 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 622..1974 /codon_start=1 /product="improved fluorescent protein-based calcium sensor (Chen et al., 2013)" /label=GCaMP6f /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK TGHAVRAIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGP VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFDKGGDGTITTKELGTVMRS LGQNPTEAELQDMINEVDAGGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK" CDS 622..1974 /codon_start=1 /product="improved fluorescent protein-based calcium sensor (Chen et al., 2013)" /label=GCaMP6s /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK TGHAVRAIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGP VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFDKGGDGTITTKELGTVMRS LGQNPTEAELQDMINEVDAGGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK" regulatory 718..727 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" primer_bind 1008..1029 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(1135..1156) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1396..1415) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" polyA_signal 2096..2217 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2133..2152) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2187..2206 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2224..2679) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2361..2382) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2573..2592 /label=F1ori-R /note="F1 origin, reverse primer" promoter 2706..2810 /gene="bla" /label=AmpR promoter promoter 2812..3169 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(2835..2855) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3020..3155 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3082..3101 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3204..3998 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(3258..3277) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3868..3887 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4186..4205) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4230..4277 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 4606..5194 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5095..5114 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg 61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca 181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc 241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta 301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac 361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg 421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg 481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt 541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta 601 ccggactcag atctcgccac catgggttct catcatcatc atcatcatgg tatggctagc 661 atgactggtg gacagcaaat gggtcgggat ctgtacgacg atgacgataa ggatctcgcc 721 accatggtcg actcatcacg tcgtaagtgg aataagacag gtcacgcagt cagagctata 781 ggtcggctga gctcactcga gaacgtctat atcaaggccg acaagcagaa gaacggcatc 841 aaggcgaact tccacatccg ccacaacatc gaggacggcg gcgtgcagct cgcctaccac 901 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg 961 agcgtgcagt ccaaactttc gaaagacccc aacgagaagc gcgatcacat ggtcctgctg 1021 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gggcggtacc 1081 ggagggagca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc 1141 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg tgagggcgat 1201 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc 1261 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac 1321 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacat ccaggagcgc 1381 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc 1441 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc 1501 ctggggcaca agctggagta caacctgccg gaccaactga ctgaagagca gatcgcagaa 1561 tttaaagagg aattctccct atttgacaag ggcggggatg ggacaataac aaccaaggag 1621 ctggggacgg tgatgcggtc tctggggcag aaccccacag aagcagagct gcaggacatg 1681 atcaatgaag tagatgccgg cggtgacggc acaatcgact tccctgagtt cctgacaatg 1741 atggcaagaa aaatgaaata cagggacacg gaagaagaaa ttagagaagc gttcggtgtg 1801 tttgataagg atggcaatgg ctacatcagt gcagcagagc ttcgccacgt gatgacaaac 1861 cttggagaga agttaacaga tgaagaggtt gatgaaatga tcagggaagc agacatcgat 1921 ggggatggtc aggtaaacta cgaagagttt gtacaaatga tgacagcgaa gtgagcggcc 1981 gcgactctag atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa 2041 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt 2101 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa 2161 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaagg 2221 cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct 2281 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg 2341 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact 2401 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac 2461 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga 2521 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga 2581 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca 2641 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa 2701 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca 2761 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg 2821 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc 2881 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc 2941 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt 3001 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc 3061 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct 3121 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag 3181 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg 3241 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg 3301 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc 3361 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga 3421 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc 3481 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag 3541 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat 3601 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg 3661 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca 3721 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct 3781 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg 3841 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg 3901 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc 3961 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat 4021 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta 4081 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg 4141 ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga 4201 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg 4261 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac 4321 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc 4381 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat 4441 agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 4501 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc 4561 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt 4621 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt 4681 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 4741 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 4801 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 4861 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 4921 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 4981 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5041 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5101 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 5161 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 5221 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 5281 tgtggataac cgtattaccg ccatgcat //