LOCUS       sequence-128553-        7641 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Light-insensitive mutant of OptoSTIM1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7641)
  AUTHORS   Kyung T, Lee S, Kim JE, Cho T, Park H, Jeong YM, Kim D, Shin A, Kim 
            S, Baek J, Kim J, Kim NY, Woo D, Chae S, Kim CH, Shin HS, Han YM, 
            Kim D, Heo WD
  TITLE     Optogenetic control of endogenous Ca channels in vivo.
  JOURNAL   Nat Biotechnol. 2015 Sep 14. doi: 10.1038/nbt.3350.
  PUBMED    26368050
REFERENCE   2  (bases 1 to 7641)
  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 
            Biotechnol. 2015 Sep 14. doi: 10.1038/nbt.3350."
COMMENT     Sequence Label: pCMV-OptoSTIM1(Cry2(D387A))
FEATURES             Location/Qualifiers
     source          1..7641
                     /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      597..606
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(603..605,606..608,609..1319)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(603..605,606..608,609..1319)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(648..669)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(909..928)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1256..1277
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             complement(2863..2880)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    4429..4550
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4466..4485)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4520..4539
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4557..5012)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4694..4715)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4906..4925
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5039..5143
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5145..5502
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5145..5336
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5168..5188)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5353..5488
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5537..6331
                     /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             5537..6331
                     /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(5591..5610)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6201..6220
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6519..6538)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6563..6610
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6939..7527
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7428..7447
                     /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 gctagcgcca
      601 ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
      661 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
      721 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
      781 ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga
      841 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
      901 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
      961 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     1021 acaagctgga gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga
     1081 acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg
     1141 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     1201 actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     1261 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagc
     1321 caccggtcgc caccatgaag atggacaaaa agaccatcgt ctggtttcgg agagatttga
     1381 gaatagaaga taatcccgcg ctcgccgccg cggcccacga gggttccgtc ttccccgttt
     1441 tcatttggtg tcctgaagaa gaaggccagt tttatcccgg aagggcctct aggtggtgga
     1501 tgaagcaaag tctggcccat cttagccagt cactgaaagc actgggcagt gatcttaccc
     1561 tgatcaagac acacaatacc atctctgcca tcctcgactg catcagggtg accggcgcaa
     1621 cgaaagtcgt gtttaaccac ctgtacgatc cagttagtct ggtgcgcgac cacactgtga
     1681 aggagaagct ggtggaacgg gggatcagtg tgcagagcta caacggggac cttctgtacg
     1741 agccatggga gatctattgc gagaaaggga aaccgttcac ctccttcaac agttactgga
     1801 agaaatgttt ggatatgtca atagagtccg ttatgttgcc ccctccctgg agactgatgc
     1861 cgattactgc tgctgcagag gccatctggg cctgctccat cgaggaactc ggtctggaaa
     1921 atgaagcaga aaagccaagc aatgcacttc tcactagagc ctggagcccc ggctggtcta
     1981 atgccgacaa gctgcttaac gagttcatcg aaaaacaact gattgactac gcgaagaact
     2041 ccaagaaagt ggtaggtaac tcaactagct tgctctctcc atatctccat tttggcgaga
     2101 tttctgtccg ccatgtattt cagtgcgctc ggatgaaaca gattatctgg gctcgcgata
     2161 aaaacagcga aggcgaagaa agcgccgatc tgttcctgcg agggatcgga cttcgggaat
     2221 actcccggta tatatgtttc aactttccat tcacacacga gcagagtctg ttgtcccacc
     2281 tcaggttctt cccctgggac gccgatgtcg acaaattcaa ggcatggaga cagggaagga
     2341 cagggtaccc actcgtggat gctggcatga gagagctctg ggctacaggc tggatgcaca
     2401 accgcatccg ggtaatcgtg tcctcatttg ctgtcaagtt tctgctcctg ccttggaaat
     2461 ggggaatgaa gtacttttgg gatacccttc tcgccgccga cttggagtgt gacattctgg
     2521 gatggcaata tattagcggg tcaattcctg acggccatga gttggacagg ttggacaatc
     2581 cggccttgca gggagctaag tatgatcccg aaggagagta tattcgacag tggctccccg
     2641 agctggcccg acttcctacg gagtggattc accatccttg ggacgcacca ctgacagtgc
     2701 tcaaggcaag cggggtggag ctgggcacca attacgctaa gcctatagtt gatatagata
     2761 cagcacgcga gctgctggct aaagcgatct ctcgcactcg ggaggcgcag attatgatcg
     2821 gtgctgccgt gtacagtggg ggaggcggag ggggtggttc aggtggtggt ggtggtggtg
     2881 gttcgggtgg tggcggaggg ggaggctccg gactcagatc taaggagcac atgaagaaga
     2941 tgatgaagga cttggagggg ttacaccgag ctgagcagag tctgcatgac cttcaggaaa
     3001 ggctgcacaa ggcccaggag gagcaccgca cagtggaggt ggagaaggtc catctggaaa
     3061 agaagctgcg cgatgagatc aaccttgcta agcaggaagc ccagcggctg aaggagctgc
     3121 gggagggtac tgagaatgag cggagccgcc aaaaatatgc tgaggaggag ttggagcagg
     3181 ttcgggaggc cttgaggaaa gcagagaagg agctagaatc tcacagctca tggtatgctc
     3241 cagaggccct tcagaagtgg ctgcagctga cacatgaggt ggaggtgcaa tattacaaca
     3301 tcaagaagca aaatgctgag aagcagctgc tggtggccaa ggagggggct gagaagataa
     3361 aaaagaagag aaacacactc tttggcacct tccacgtggc ccacagctct tccctggatg
     3421 atgtagatca taaaattcta acagctaagc aagcactgag cgaggtgaca gcagcattgc
     3481 gggagcgcct gcaccgctgg caacagatcg agatcctctg tggcttccag attgtcaaca
     3541 accctggcat ccactcactg gtggctgccc tcaacataga ccccagctgg atgggcagta
     3601 cacgccccaa ccctgctcac ttcatcatga ctgacgacgt ggatgacatg gatgaggaga
     3661 ttgtgtctcc cttgtccatg cagtccccta gcctgcagag cagtgttcgg cagcgcctga
     3721 cggagccaca gcatggcctg ggatctcaga gagatttgac ccattccgat tcggagtcct
     3781 ccctccacat gagtgaccgc cagcgtgtgg cccccaaacc tcctcagatg agccgtgctg
     3841 cagacgaggc tctcaatgcc atgacttcca atggcagcca ccggctgatc gagggggtcc
     3901 acccagggtc tctggtggag aaactgcctg acagccctgc cctggccaag aaggcattac
     3961 tggcgctgaa ccatgggctg gacaaggccc acagcctgat ggagctgagc ccctcagccc
     4021 cacctggtgg ctctccacat ttggattctt cccgttctca cagccccagc tccccagacc
     4081 cagacacacc atctccagtt ggggacagcc gagccctgca agccagccga aacacacgca
     4141 ttccccacct ggctggcaag aaggctgtgg ctgaggagga taatggctct attggcgagg
     4201 aaacagactc cagcccaggc cggaagaagt ttcccctcaa aatctttaag aagcctctta
     4261 agaagtagga attctgcagt cgacggtacc gcgggcccgg gatccaccgg atctagataa
     4321 ctgatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
     4381 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
     4441 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     4501 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat
     4561 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     4621 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     4681 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     4741 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     4801 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     4861 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     4921 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     4981 cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg
     5041 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     5101 ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac
     5161 cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     5221 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     5281 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     5341 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     5401 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     5461 aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg
     5521 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     5581 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     5641 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     5701 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     5761 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     5821 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     5881 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     5941 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     6001 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     6061 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     6121 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     6181 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     6241 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     6301 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     6361 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     6421 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     6481 atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata
     6541 ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg
     6601 tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg
     6661 agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt
     6721 tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca
     6781 ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     6841 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     6901 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     6961 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     7021 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     7081 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     7141 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     7201 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     7261 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     7321 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     7381 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     7441 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     7501 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     7561 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     7621 aaccgtatta ccgccatgca t
//
