LOCUS       sequence-133662-        7461 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of hBACCS2, a human blue light-activated Ca2+ 
            channel switch, and mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7461)
  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 7461)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2015-08-18"; volume: "6:8021. doi"; pages: " 
            10.1038/ncomms9021"
COMMENT     Sequence Label: hBACCS2-IRES-mCherry
FEATURES             Location/Qualifiers
     source          1..7461
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     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)"
     primer_bind     complement(2353..2370)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2580..2599
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2761..3471
                     /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(2906..2924)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3174..3193
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    3516..4104
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3569..3589)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3987..3998)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4249..4370
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4286..4305)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4340..4359
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4377..4832)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4514..4535)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4726..4745
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4859..4963
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4965..5322
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4965..5156
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4988..5008)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5173..5308
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5357..6151
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             5357..6151
                     /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(5411..5430)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6021..6040
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6339..6358)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6383..6430
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6759..7347
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7248..7267
                     /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 aatcttcaga agtggctgca gctgacacat gaggtggagg
     1081 tgcaatatta caacatcaag aagcaaaatg ctgagaagca gctgctggtg gccaaggagg
     1141 gggctgagaa gataaaaaag aagagaaaca cactctttgg caccttccac gtggcccaca
     1201 gctcttccct ggatgatgta gatcataaaa ttctaacagc taagcaagca ctgagcgagg
     1261 tgacagcagc attgcgggag cgcctgcacc gctggcaaca gatcgagatc ctctgtggct
     1321 tccagattgt caacaaccct ggcatccacg 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 ttcagaagtg gctgcagctg acacatgagg
     1861 tggaggtgca atattacaac atcaagaagc aaaatgctga gaagcagctg ctggtggcca
     1921 aggagggggc tgagaagata aaaaagaaga gaaacacact ctttggcacc ttccacgtgg
     1981 cccacagctc ttccctggat gatgtagatc ataaaattct aacagctaag caagcactga
     2041 gcgaggtgac agcagcattg cgggagcgcc tgcaccgctg gcaacagatc gagatcctct
     2101 gtggcttcca gattgtcaac aaccctggca tccacggcgg 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 atggtgagca agggcgagga ggataacatg gccatcatca aggagttcat gcgcttcaag
     2821 gtgcacatgg agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc
     2881 cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc
     2941 ttcgcctggg acatcctgtc ccctcagttc atgtacggct ccaaggccta cgtgaagcac
     3001 cccgccgaca tccccgacta cttgaagctg tccttccccg agggcttcaa gtgggagcgc
     3061 gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc cctgcaggac
     3121 ggcgagttca tctacaaggt gaagctgcgc ggcaccaact tcccctccga cggccccgta
     3181 atgcagaaga agaccatggg ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc
     3241 gccctgaagg gcgagatcaa gcagaggctg aagctgaagg acggcggcca ctacgacgct
     3301 gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc
     3361 aacatcaagt tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgaa
     3421 cgcgccgagg gccgccactc caccggcggc atggacgagc tgtacaagta accgcggtcg
     3481 acggtaccgc gggcccggga tctaagctta tcgataatca acctctggat tacaaaattt
     3541 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     3601 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     3661 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     3721 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     3781 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     3841 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     3901 tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc
     3961 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     4021 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
     4081 tctccctttg ggccgcctcc ccgcatcgat accgtggatc gatccaccgg atctagataa
     4141 ctgatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
     4201 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
     4261 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     4321 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat
     4381 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     4441 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     4501 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     4561 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     4621 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     4681 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     4741 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     4801 cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg
     4861 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     4921 ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac
     4981 cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     5041 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     5101 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     5161 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     5221 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     5281 aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg
     5341 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     5401 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     5461 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     5521 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     5581 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     5641 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     5701 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     5761 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     5821 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     5881 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     5941 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     6001 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     6061 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     6121 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     6181 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     6241 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     6301 atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata
     6361 ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg
     6421 tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg
     6481 agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt
     6541 tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca
     6601 ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     6661 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     6721 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     6781 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     6841 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     6901 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     6961 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     7021 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     7081 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     7141 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     7201 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     7261 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     7321 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     7381 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     7441 aaccgtatta ccgccatgca t
//
