LOCUS       sequence-133661-        7470 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of hBACCS2, a human blue light-activated Ca2+ 
            channel switch, and GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7470)
  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 7470)
  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-GFP
FEATURES             Location/Qualifiers
     source          1..7470
                     /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             join(2761..2763,2764..2766,2767..3480)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2761..2763,2764..2766,2767..3477)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2806..2827)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3067..3086)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3414..3435
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3525..4113
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3578..3598)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3996..4007)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4258..4379
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4295..4314)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4349..4368
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4386..4841)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4523..4544)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4735..4754
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4868..4972
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4974..5331
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4974..5165
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4997..5017)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5182..5317
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5366..6160
                     /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             5366..6160
                     /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(5420..5439)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6030..6049
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6348..6367)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6392..6439
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6768..7356
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7257..7276
                     /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 gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     2821 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     2881 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     2941 ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag
     3001 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     3061 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     3121 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     3181 aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac
     3241 ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
     3301 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     3361 tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     3421 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa
     3481 ccgcggtcga cggtaccgcg ggcccgggat ctaagcttat cgataatcaa cctctggatt
     3541 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     3601 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     3661 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
     3721 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
     3781 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
     3841 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     3901 ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt gttgccacct
     3961 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
     4021 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
     4081 cgagtcggat ctccctttgg gccgcctccc cgcatcgata ccgtggatcg atccaccgga
     4141 tctagataac tgatcataat cagccatacc acatttgtag aggttttact tgctttaaaa
     4201 aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac
     4261 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     4321 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa
     4381 cgcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     4441 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     4501 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     4561 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     4621 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     4681 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     4741 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     4801 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca cttttcgggg
     4861 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     4921 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtcctgaggc
     4981 ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     5041 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     5101 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     5161 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     5221 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     5281 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa agatcgatca
     5341 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     5401 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     5461 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     5521 cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac
     5581 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     5641 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     5701 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     5761 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     5821 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     5881 caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     5941 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     6001 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     6061 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     6121 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa
     6181 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     6241 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     6301 ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga aacacggaag
     6361 gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa taaaacgcac
     6421 ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca ctctgtcgat
     6481 accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc cccaccccac
     6541 cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc aggccctgcc
     6601 atagcctcag gttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     6661 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     6721 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     6781 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     6841 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     6901 ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     6961 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     7021 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     7081 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     7141 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     7201 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     7261 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     7321 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     7381 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     7441 tctgtggata accgtattac cgccatgcat
//
