LOCUS Exported 6625 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Ratiometric (Blue to Green) fluorescent indicator for calcium signaling in the endoplasmic reticulum. ACCESSION . VERSION . KEYWORDS pCIS GEM-CEPIA1er SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6625) AUTHORS Suzuki J, Kanemaru K, Ishii K, Ohkura M, Okubo Y, Iino M TITLE Imaging intraorganellar Ca(2+) at subcellular resolution using CEPIA. JOURNAL Nat Commun. 2014 Jun 13;5:4153. doi: 10.1038/ncomms5153. PUBMED 24923787 REFERENCE 2 (bases 1 to 6625) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6625 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin 4..459 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(91..110) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 301..322 /label=F1ori-F /note="F1 origin, forward primer" primer_bind 585..607 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 599..616 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 600..616 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 626..645 /label=T7 /note="T7 promoter, forward primer" promoter 626..644 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" enhancer 709..1012 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 1015..1290 /label=chicken beta-actin promoter intron 1291..2299 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind 2223..2245 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 2355..2371 /label=pBluescriptKS /note="For pBluescript vector" primer_bind 2356..2372 /label=KS primer /note="common sequencing primer, one of multiple similar variants" regulatory 2381..2390 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" sig_peptide join(2387..2431,2514..2525) /product="modified signal sequence from the mouse immunoglobulin heavy chain variable region" /label=IgH signal sequence CDS 2562..2618 /codon_start=1 /product="calmodulin-binding peptide from avian smooth muscle myosin light chain kinase" /label=calmodulin-binding peptide /translation="RRKWNKTGHAVRAIGRLSS" primer_bind 2831..2852 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(3216..3235) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" CDS 3801..3830 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" primer_bind complement(3964..3983) /label=Bglob-pA-R /note="Rabbit beta-globin polyA region, reverse primer" polyA_signal 4029..4084 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind complement(4083..4102) /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" primer_bind complement(4437..4457) /label=T3 /note="T3 promoter, forward primer" promoter complement(4437..4455) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(4476..4492) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4476..4492) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(4489..4511) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 4500..4516 /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)." promoter complement(4524..4554) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4569..4590 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(4707..4724) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4878..5466) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4958..4977) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5637..6497) /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 6260..6279 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6498..6602) /gene="bla" /label=AmpR promoter ORIGIN 1 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga 61 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg 121 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat 181 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg 241 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata 301 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt 361 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat 421 ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc cattcgccat tcaggctgcg 481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg 541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg 601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtaccta 661 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact 721 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat 781 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggacta 841 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc 901 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg 961 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tgggtcgagg 1021 tgagccccac gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt 1081 atttatttat tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggcgcgcg 1141 ccaggcgggg cggggcgggg cgaggggcgg ggcggggcga ggcggagagg tgcggcggca 1201 gccaatcaga gcggcgcgct ccgaaagttt ccttttatgg cgaggcggcg gcggcggcgg 1261 ccctataaaa agcgaagcgc gcggcgggcg ggagtcgctg cgttgccttc gccccgtgcc 1321 ccgctccgcg ccgcctcgcg ccgcccgccc cggctctgac tgaccgcgtt actcccacag 1381 gtgagcgggc gggacggccc ttctcctccg ggctgtaatt agcgcttggt ttaatgacgg 1441 ctcgtttctt ttctgtggct gcgtgaaagc cttaaagggc tccgggaggg ccctttgtgc 1501 gggggggagc ggctcggggg gtgcgtgcgt gtgtgtgtgc gtggggagcg ccgcgtgcgg 1561 cccgcgctgc ccggcggctg tgagcgctgc gggcgcggcg cggggctttg tgcgctccgc 1621 gtgtgcgcga ggggagcgcg gccgggggcg gtgccccgcg gtgcgggggg gctgcgaggg 1681 gaacaaaggc tgcgtgcggg gtgtgtgcgt gggggggtga gcagggggtg tgggcgcggc 1741 ggtcgggctg taaccccccc ctgcaccccc ctccccgagt tgctgagcac ggcccggctt 1801 cgggtgcggg gctccgtgcg gggcgtggcg cggggctcgc cgtgccgggc ggggggtggc 1861 ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc cggggagggc tcgggggagg 1921 ggcgcggcgg ccccggagcg ccggcggctg tcgaggcgcg gcgagccgca gccattgcct 1981 tttatggtaa tcgtgcgaga gggcgcaggg acttcctttg tcccaaatct ggcggagccg 2041 aaatctggga ggcgccgccg caccccctct agcgggcgcg ggcgaagcgg tgcggcgccg 2101 gcaggaagga aatgggcggg gagggccttc gtgcgtcgcc gcgccgccgt ccccttctcc 2161 atctccagcc tcggggctgc cgcaggggga cggctgcctt cgggggggac ggggcagggc 2221 ggggttcggc ttctggcgtg tgaccggcgg ctctagagcc tctgctaacc atgttcatgc 2281 cttcttcttt ttcctacagc tcctgggcaa cgtgctggtt gttgtgctgt ctcatcattt 2341 tggcaaagga ttcctcgagg tcgacggtat cgataagctt gccaccatgg gatggagctg 2401 tatcatcctc ttcttggtag caacagctac aggtaagggg ttaacagtag caggcttgag 2461 gtctggacat atatatgggt gacaatgaca tccactttgc ctttctctcc acaggcgcgc 2521 actcccaggt ccaactgcag ggatccatgg tcgactcttc acgtcgtaag tggaataaga 2581 caggtcacgc agtcagagct ataggtcggc tgagctcacc agagaacgtg tatataaagg 2641 ccgacgagca gaagaacggc atcaaggcgt acttcaagat ccgccacaac atcgagggcg 2701 gcggcgtgca gctcgcctac cactaccagc agatcacccc cataggcgac ggccccgtgc 2761 tgctgcccga caaccactac ctgagcgtgc agtccatact ttcgaaagac cccaacgaga 2821 agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg 2881 acgagctgta caagggtggc agcggtggca tggtgagcaa gggcgaggag ctgttcaccg 2941 gggtggtgcc catccaggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 3001 ccggcgaggg tgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca 3061 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgtcctac ggcgtgcagt 3121 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 3181 aaggctacat ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 3241 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 3301 tcaaggagga cggcaacatc ctggggcaca agctggagta cagcacgcgt gaccaactga 3361 ctgaagagca gatcgcagaa tttaaagagg ctttctccct atttgacaag gacggggatg 3421 ggacgataac aaccaaggat ctggggacgg tgatgcggtc tctggggcag aaccccacag 3481 aagcagagct ccaggacatg atcaatgaag tagatgccga cggtgacggc acaatcgact 3541 tccctgagtt cctgacaatg atggcaccta aaatgcagga cacagacagt gaagaggaaa 3601 ttcgcgaagc gttccgtgtg tgggataagg atggcaatgg ctacatcggc gcagcagagc 3661 ttcgccacgt gatgacaaac cttggagaga agttaacaga tgaagaggtt gatgaaatga 3721 tcagggtagc agacatcgat ggagaaggtc aggtaaacta cgaagacttt gtacaaatga 3781 tgacagcgaa ggcggccgca gaacaaaaac tcatctcaga agaggatctg aatggggccg 3841 caagcgagaa ggacgagctg tagactagtt ctagagcggc cgcttcactc ctcaggtgca 3901 ggctgcctat cagaaggtgg tggctggtgt ggccaatgcc ctggctcaca aataccactg 3961 agatcttttt ccctctgcca aaaattatgg ggacatcatg aagccccttg agcatctgac 4021 ttctggctaa taaaggaaat ttattttcat tgcaatagtg tgttggaatt ttttgtgtct 4081 ctcactcgga aggacatatg ggagggcaaa tcatttaaaa catcagaatg agtatttggt 4141 ttagagtttg gcaacatatg ccatatgctg gctgccatga acaaaggtgg ctataaagag 4201 gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga aaagccttga 4261 cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta acatccctaa 4321 aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta ctcccagtca 4381 tagctgtccc tcttctctta tgaagatccc tcgacctgga gctccagctt ttgttccctt 4441 tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat 4501 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg 4561 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag 4621 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt 4681 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg 4741 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg 4801 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag 4861 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga 4921 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct 4981 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc 5041 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg 5101 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc 5161 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca 5221 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag 5281 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct 5341 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc 5401 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga 5461 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca 5521 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat 5581 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac 5641 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt 5701 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt 5761 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag 5821 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct 5881 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt 5941 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc 6001 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt 6061 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg 6121 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg 6181 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct 6241 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc 6301 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt 6361 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt 6421 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg 6481 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat 6541 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg 6601 cgcacatttc cccgaaaagt gccac //