LOCUS       Exported                9118 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Mammalian expression of dmBACCS2, a Drosophila melanogaster blue 
            light-activated Ca2+ channel switch, along with dOrai and mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    dmBACCS2-IRES-dOrai-IRES-mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9118)
  AUTHORS   Ishii T, Sato K, Kakumoto T, Miura S, Touhara K, Takeuchi S, Nakata 
            T
  TITLE     Light generation of intracellular Ca(2+) signals by a genetically 
            encoded protein BACCS.
  JOURNAL   Nat Commun. 2015 Aug 18;6:8021. doi: 10.1038/ncomms9021.
  PUBMED    26282514
REFERENCE   2  (bases 1 to 9118)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9118
                     /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      603..612
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     misc_feature    2186..2759
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    2209..2761
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     regulatory      3281..3290
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     misc_feature    3843..4416
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    3866..4418
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     CDS             4418..5128
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4563..4581)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4831..4850
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    5173..5761
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5226..5246)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5644..5655)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    5906..6027
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5943..5962)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5997..6016
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(6034..6489)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6171..6192)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6383..6402
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6516..6620
                     /gene="bla"
                     /label=AmpR promoter
     promoter        6622..6979
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     primer_bind     complement(6645..6665)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6830..6965
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6892..6911
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             7014..7808
                     /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(7068..7087)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7678..7697
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7996..8015)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    8040..8087
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      8416..9004
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8905..8924
                     /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 gctagcctcg
      601 aggccaccat gggaccggtc ggaggaagcg gtggatcttt ggctactaca cttgaacgta
      661 ttgagaagaa ctttgtcatt actgacccaa gattgccaga taatcccatt atattcgcgt
      721 ccgatagttt cttgcagttg acagaatata gccgtgaaga aattttggga agaaactgca
      781 ggtttctaca aggtcctgaa actgatcgcg cgacagtgag aaaaattaga gatgccatag
      841 ataaccaaac agaggtcact gttcagctga ttaattatac aaagagtggt aaaaagttct
      901 ggaacctctt tcacttgcag cctatgcgag atcagaaggg agatgtccag tactttattg
      961 gggttcagtt ggatggaact gagcatgtcc gagatgctgc cgagagagag ggagtcatgc
     1021 tgattaagaa aactgcagaa aatctgcaat catggcttca atacacatat gaactagaaa
     1081 gtaagaatca tcagaagaag cgcacgtcgg ctgagaagca gctacagtcg gccagagagg
     1141 cttgtgagaa attgcgtaag aaacggtcaa gtttggtggg tgcgttcgtt tccacgcacg
     1201 gaaagagtat tgatgatgtg gatcggtcga ttgttgaggc acggaatgcc ctcggagatg
     1261 taacaaacga gctgcaagaa cgactgcatc gctggaagca aatcgagacg tgccttggct
     1321 taaacattgt gaacaacaat ggtctgcccg gaggatccgg aggaagtgga ggaggtatcc
     1381 tgcagtctcg gggtggcagt ggtggttcag gcggttctgg atctttggct actacacttg
     1441 aacgtattga gaagaacttt gtcattactg acccaagatt gccagataat cccattatat
     1501 tcgcgtccga tagtttcttg cagttgacag aatatagccg tgaagaaatt ttgggaagaa
     1561 actgcaggtt tctacaaggt cctgaaactg atcgcgcgac agtgagaaaa attagagatg
     1621 ccatagataa ccaaacagag gtcactgttc agctgattaa ttatacaaag agtggtaaaa
     1681 agttctggaa cctctttcac ttgcagccta tgcgagatca gaagggagat gtccagtact
     1741 ttattggggt tcagttggat ggaactgagc atgtccgaga tgctgccgag agagagggag
     1801 tcatgctgat taagaaaact gcagaaaatc tgcaatcatg gcttcaatac acatatgaac
     1861 tagaaagtaa gaatcatcag aagaagcgca cgtcggctga gaagcagcta cagtcggcca
     1921 gagaggcttg tgagaaattg cgtaagaaac ggtcaagttt ggtgggtgcg ttcgtttcca
     1981 cgcacggaaa gagtattgat gatgtggatc ggtcgattgt tgaggcacgg aatgccctcg
     2041 gagatgtaac aaacgagctg caagaacgac tgcatcgctg gaagcaaatc gagacgtgcc
     2101 ttggcttaaa cattgtgaac aacaatggtc tgcccggcgg ctctggaggt agcggactag
     2161 tttgagtcga gccgcggcaa ttccgcccct ctccctcccc cccccctaac gttactggcc
     2221 gaagccgctt ggaataaggc cggtgtgcgt ttgtctatat gttattttcc accatattgc
     2281 cgtcttttgg caatgtgagg gcccggaaac ctggccctgt cttcttgacg agcattccta
     2341 ggggtctttc ccctctcgcc aaaggaatgc aaggtctgtt gaatgtcgtg aaggaagcag
     2401 ttcctctgga agcttcttga agacaaacaa cgtctgtagc gaccctttgc aggcagcgga
     2461 accccccacc tggcaacagg tgcctctgcg gccaaaagcc acgtgtataa gatacacctg
     2521 caaaggcggc acaaccccag tgccacgttg tgagttggat agttgtggaa agagtcaaat
     2581 ggctctcctc aagcgtattc aacaaggggc tgaaggatgc ccagaaggta ccccattgta
     2641 tgggatctga tctggggcct cggtgcacat gctttacatg tgtttagtcg aggttaaaaa
     2701 aacgtctagg ccccccgaac cacggggacg tggttttcct ttgaaaaaca cgatgataat
     2761 atgtctgtgt ggaccacggc caacaattcg ggcctagaga cgccaacaaa gtcgccgatc
     2821 acgtcgtcgg ttccgcgggc atcgagaagt tctgcggtga tcaccactgg aaatcaccag
     2881 cagcaccact tccagcacgt tgtggccgcc gccgttgcag ccgccacttc agtggccacc
     2941 ggtcatcagt tccagcagca gttcccgctg cacgcacatc cgcatccgca tccacagcac
     3001 cacagcaaca gtcccacggg tagcggcagc aacagcaaca acagcgcagg tttccagcgc
     3061 accagcatca gcaactcgct gctgcagttc ccgccgcccc cgcccccttc ctcacagaac
     3121 caggcgaagc cgcgcggaca ccatcgcacc gccagcagca gcatgtcgca atccggcgag
     3181 gatctccact cgcccaccta cctgtcctgg cggaagctcc agcttagtcg ggccaagcta
     3241 aaggcatcca gcaagacgtc cgccctgctc tccggattcg ccatggtggc gatggtggag
     3301 gtgcaactgg atcacgatac gaatgtacca ccgggcatgc tgatagcctt cgccatctgc
     3361 accaccctcc tggtggcggt gcacatgctg gccctgatga tcagcacctg tatcctgcca
     3421 aatattgaga cggtgtgtaa tttgcatagt atttccctgg tccacgaatc gccacacgag
     3481 cgtctccatt ggtacattga aacggcgtgg gcattctcca cgctgctggg actcatactc
     3541 tttctactgg agatagccat cctgtgctgg gtgaaattct atgacctaag tccgccggcg
     3601 gcttggtcag cgtgtgtggt cctcataccg gtgatgatca tctttatggc cttcgccatt
     3661 cacttctatc gctccctggt gtcgcacaaa tatgaggtga cagtctcggg catccgtgag
     3721 ctggagatgc tcaaggagca gatggagcag gatcatttgg agcatcacaa taacatacga
     3781 aataatggca tgaactatgg cgcatccggg gacattgtct aggtcgagcc gcggcaattc
     3841 cgcccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga ataaggccgg
     3901 tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa tgtgagggcc
     3961 cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc tctcgccaaa
     4021 ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc ttcttgaaga
     4081 caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg caacaggtgc
     4141 ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca accccagtgc
     4201 cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag cgtattcaac
     4261 aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct ggggcctcgg
     4321 tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc cccgaaccac
     4381 ggggacgtgg ttttcctttg aaaaacacga tgataatatg gtgagcaagg gcgaggagga
     4441 taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa
     4501 cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac
     4561 cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc
     4621 tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt
     4681 gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg
     4741 cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa
     4801 gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga ccatgggctg
     4861 ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca
     4921 gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc
     4981 caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc
     5041 ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac
     5101 cggcggcatg gacgagctgt acaagtaacc gcggtcgacg gtaccgcggg cccgggatct
     5161 aagcttatcg ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     5221 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     5281 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt
     5341 tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac
     5401 gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct
     5461 ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca
     5521 ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt
     5581 ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc
     5641 ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct
     5701 cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg
     5761 catcgatacc gtggatcgat ccaccggatc tagataactg atcataatca gccataccac
     5821 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
     5881 taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata
     5941 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     6001 tttgtccaaa ctcatcaatg tatcttaacg cgtaaattgt aagcgttaat attttgttaa
     6061 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     6121 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     6181 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     6241 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     6301 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     6361 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     6421 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     6481 agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     6541 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     6601 aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt
     6661 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     6721 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     6781 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     6841 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     6901 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     6961 aggcctaggc ttttgcaaag atcgatcaag agacaggatg aggatcgttt cgcatgattg
     7021 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     7081 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     7141 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg
     7201 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     7261 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     7321 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     7381 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     7441 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     7501 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg
     7561 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     7621 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     7681 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     7741 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     7801 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     7861 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     7921 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     7981 tagggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac
     8041 ggcaataaaa agacagaata aaacgcacgg tgttgggtcg tttgttcata aacgcggggt
     8101 tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc
     8161 ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc
     8221 agccaacgtc ggggcggcag gccctgccat agcctcaggt tactcatata tactttagat
     8281 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     8341 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     8401 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     8461 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     8521 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
     8581 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     8641 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     8701 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     8761 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     8821 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     8881 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     8941 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     9001 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     9061 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg ccatgcat
//
