LOCUS       Exported                5341 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expresses red fluorescent calcium sensor in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pGP-CMV-NES-fRCaMP2
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5341)
  AUTHORS   Kerruth S, Coates C, Durst CD, Oertner TG, Torok K
  TITLE     The kinetic mechanisms of fast-decay red-fluorescent genetically 
            encoded calcium indicators.
  JOURNAL   J Biol Chem. 2019 Mar 15;294(11):3934-3946. doi: 
            10.1074/jbc.RA118.004543. Epub 2019 Jan 16.
  PUBMED    30651353
REFERENCE   2  (bases 1 to 5341)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5341
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        75..378
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        379..582
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     533..553
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             702..719
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             723..755
                     /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             759..782
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             810..866
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKTGHAVRAIGRLSS"
     polyA_signal    2143..2264
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2180..2199)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2234..2253
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2271..2726)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2408..2429)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2620..2639
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2753..2857
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2859..3216
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(2882..2902)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3067..3202
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3129..3148
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3251..4045
                     /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(3305..3324)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3915..3934
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4233..4252)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4277..4324
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4653..5241
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5142..5161
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttaccgccat gcattagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
       61 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
      121 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      181 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      241 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      301 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      361 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      421 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      481 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      541 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag
      601 atccgctagc gctaccggac tcagatctcg ccaccatgct gcagaacgag cttgctctta
      661 agttggctgg acttgatatt aacaagactg gaggaggttc tcatcatcat catcatcatg
      721 gtatggctag catgactggt ggacagcaaa tgggtcggga tctgtacgac gatgacgata
      781 aggatctcgc aacaatggtc gactcatcac gtcgtaagtg gaataagaca ggtcacgcag
      841 tcagagctat aggtcggctg agctcagcga tcaactgcga aatgatgtac ccagcggatg
      901 gtggtctgcg tggttacact cacatggcgc tgaaagttga tggcggcggt cacctgtcct
      961 gttctttcgt gaccacctac cgctccaaaa agactgtcgg caacattaag atgcctgcca
     1021 ttcatagcgt cagccaccgt ctggagcgcc tggaggagag cgataacgaa atgtttgtcg
     1081 tacagcgtga acacgcagtt gccaagtttg tgggcctggg tggtggcggc ggtaccggag
     1141 ggagcatgaa ctccctgatc aaggagaaca tgcgtatgaa agtggttctg gaaggctccg
     1201 taaacggcca ccagttcaaa tgcactggtg aaggcgaagg caacccgtat atgggcaccc
     1261 agactatgcg tatcaaagtg atcgagggtg gtccgctgcc gtttgcgttc gacatcctgg
     1321 cgacgtcctt tatgtatggc tcccgtacct tcatcaaata tccgaaaggc atcccggatt
     1381 tctttaagca gtccttcccg gaaggtttta cctgggaacg tgtgacccgt tacgaagacg
     1441 gcggcgtaat taccgttatg caagacacgt ctctggagga tggctgcctg gtgtatcacg
     1501 tgcaggttcg cggtgtgaac ttcccgagca atggtgctgt aatgcaaaag aaaaccaaag
     1561 gttgggagcc tacggactcc caaaccactg aagagcagat cgcagaattt aaagaggctt
     1621 tctccctatt tgacaaggac ggggatggga caataacaac caaggagctg gggacggtga
     1681 tgcggtctct ggggcagaac cccacagaag cagagctgca ggacatgatc aatgaagtag
     1741 atgccgacgg tgacggcaca atcgacttcc ctgagttcct gattatgatg gcaagaaaaa
     1801 tgaaatacac agacagtgaa gaagaaatta gagaagcgtt cggcgtgttt gctaaggatg
     1861 gcaatggcta catcagtgca gcagagcttc gccacgtgat gacaaacctt ggagagaagt
     1921 taacagatga agaggttgat gaaatgatca gggaagcaga cagcgatggg gatggtcagg
     1981 taaactacga agagtttgta caaatgatga cagcgaagta ggcggccgcg actctagatc
     2041 ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc
     2101 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct
     2161 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     2221 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt aaattgtaag
     2281 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     2341 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     2401 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     2461 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     2521 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     2581 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     2641 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     2701 gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac
     2761 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     2821 ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg
     2881 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     2941 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     3001 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     3061 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     3121 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     3181 tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg
     3241 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     3301 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     3361 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     3421 gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     3481 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     3541 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     3601 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     3661 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     3721 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag
     3781 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     3841 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     3901 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     3961 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     4021 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     4081 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     4141 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     4201 gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa
     4261 ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt
     4321 gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc
     4381 cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt
     4441 gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac
     4501 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     4561 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     4621 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     4681 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     4741 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     4801 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     4861 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     4921 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     4981 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     5041 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     5101 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     5161 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     5221 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     5281 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     5341 a
//
