pcDNA-K-GECO1

LOCUS       pcDNA-K-GECO1                6598 bp ds-DNA     circular SYN 06-AUG-2018
DEFINITION  Mammalian cell expression plasmid for red fluorescent Ca2+ indicator
            K-GECO1.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA-K-GECO1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6598)
  AUTHORS   Shen Y, Dana H, Abdelfattah AS, Patel R, Shea J, Molina RS, Rawal B,
            Rancic V, Chang YF, Wu L, Chen Y, Qian Y, Wiens MD, Hambleton N, 
            Ballanyi K, Hughes TE, Drobizhev M, Kim DS, Koyama M, Schreiter ER, 
            Campbell RE
  TITLE     A genetically encoded Ca(2+) indicator based on circularly 
            permutated sea anemone red fluorescent protein eqFP578.
  JOURNAL   BMC Biol. 2018 Jan 16;16(1):9. doi: 10.1186/s12915-018-0480-0.
  PUBMED    29338710
REFERENCE   2  (bases 1 to 6598)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 6, 2018 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6598
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        110..489
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        490..693
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     644..664
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     738..757
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        738..756
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      781..790
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     primer_bind     complement(2064..2081)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2070..2294
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2340..2768
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2427..2446)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2637..2658
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2777..2797)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2782..3111
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2962..3097
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3024..3043
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3178..3972
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3232..3251)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3842..3861
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4146..4267
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4183..4202)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4237..4256
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4316..4332)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4316..4332)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4329..4351)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4340..4356
                     /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(4364..4394)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4409..4430
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4547..4564)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4718..5306)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4798..4817)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5477..6337)
                     /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     6100..6119
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6338..6442)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6517..6536)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
       61 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
      121 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
      181 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      241 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      301 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      361 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      421 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      481 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      541 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      601 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      661 cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga acccactgct
      721 tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg ctagcgtgcc
      781 gccaccatgg cctcgagaat gggcagcgtg aagctgatcc ccagcctgac caccgtgatc
      841 ctcgtgaagt ccatgctgcg gaagcggagc ttcggcaacc ccttcaagta taatacggag
      901 accctgtacc ccgctgacgg cggcctggaa ggcgcatgtg acatggccct gaagctcgtg
      961 ggcgggggcc acctgaactg cagccttgag accacataca gatccaagaa acccgctacg
     1021 aacctcaaga tgcccggtgt ctacaacgtg gaccacagac tggaacgaat caaagaggcc
     1081 gacgatgaga cctacgtcga gctgcacgag gtggctgtgg ccagatacgt gggcctgggt
     1141 ggtggcggag gtacaggcgg gagtgtgagc gagctgatta aggagaacat gccaatgaag
     1201 ctgtacatgg agggcaccgt gaacaaccac cacttcaagt gcacatccga gggcgaaggc
     1261 aagccctacg agggcaccca gaccatgaga atcaaggtcg tcgagggcgg ccctctcccc
     1321 ttcgccttcg acatcctggc taccagcttc atgtacggca gcagaacctt catcaagcac
     1381 cctcctggca tccccgactt ctttaagcag tccttccctg agggcttcac atgggagaga
     1441 gtcaccacat acgaagacgg gggcgtgctg accgctaccc aggacaccag cctccaggac
     1501 ggctgcctca tctacaacgt caaggttaga gggatgaact tcccagccaa cggccctgtg
     1561 atgcagaaga aaacactcgg ctgggaggcc agtaatggcc aactgactga agagcagatc
     1621 gcagaattta aagaggcttt ctccctattt gacaaggacg gggatgggac gataacaacc
     1681 aaggagctgg ggacggtgat gcggtctctg gggcagaacc ccacagaagc ggagctgcag
     1741 gacatgatca atgaagtaga tgccgacggt gacggcacat tcgacttccc tgagttcctg
     1801 acgatgatgg caagaaaaat gagttataga gtcactgaag aggaaattag agaagcgttc
     1861 cgcgtgtttg ataaggacgg caatggctac atcggcgcag cagagcttcg ccacgtgatg
     1921 acagaccttg gagagaagtt aacagatgag gaggttgatg aaatgatcag ggtagcagac
     1981 atcgatgggg atggtcaggt aaactacgaa gagtttgtac aaatgatgac agcgaagtag
     2041 aagcttggta aaccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     2101 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     2161 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     2221 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     2281 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
     2341 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2401 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2461 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2521 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2581 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     2641 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     2701 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     2761 cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     2821 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     2881 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     2941 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     3001 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag
     3061 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg
     3121 ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg
     3181 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3241 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     3301 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     3361 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     3421 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     3481 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     3541 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     3601 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     3661 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     3721 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     3781 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     3841 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     3901 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     3961 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
     4021 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     4081 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     4141 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     4201 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     4261 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     4321 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     4381 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     4441 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4501 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     4561 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     4621 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     4681 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     4741 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     4801 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     4861 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     4921 gctataggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     4981 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     5041 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     5101 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     5161 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     5221 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     5281 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5341 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5401 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5461 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     5521 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5581 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5641 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5701 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     5761 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     5821 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     5881 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     5941 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6001 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6061 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     6121 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6181 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6241 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6301 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6361 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6421 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     6481 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
     6541 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagc
//