LOCUS       sequence-319200-        8008 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses LightR-bRaf-mVenus in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8008)
  AUTHORS   Shaaya M, Fauser J, Zhurikhina A, Conage-Pough JE, Huyot V, Brennan 
            M, Flower CT, Matsche J, Khan S, Natarajan V, Rehman J, Kota P, 
            White FM, Tsygankov D, Karginov AV
  TITLE     Light-Regulated allosteric switch enables temporal and subcellular 
            control of enzyme activity.
  JOURNAL   Elife. 2020 Sep 23;9. pii: 60647. doi: 10.7554/eLife.60647.
  PUBMED    32965214
REFERENCE   2  (bases 1 to 8008)
  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: "Elife."; 
            date: "2020-09-23"; volume: "9. pii: 60647. doi"; pages: " 
            10.7554/eLife.60647"
COMMENT     Sequence Label: LightR-bRaf-mVenus
FEATURES             Location/Qualifiers
     source          1..8008
                     /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"
     CDS             2562..3011
                     /codon_start=1
                     /product="""positive Magnet"" variant of an N-terminally 
                     truncated Vivid (VVD) blue light photoreceptor from 
                     Neurospora crassa (Kawano et al., 2015)"
                     /label=pMag
                     /note="rapidly heterodimerizes with nMagHigh1 upon blue 
                     light irradiation to form a complex with slow switch-off 
                     kinetics"
                     /translation="HTLYAPGGYDIMGYLIQIMNRPNPQVELGPVDTSCALILCDLKQK
                     DTPIVYASEAFLYMTGYSNAEVLGRNCRFLQSPDGMVKPKSTRKYVDSNTINTMRKAID
                     RNAEVQVEVVNFKKNGQRFVNFLTMIPVRDETGEYRYSMGFQCETE"
     regulatory      3948..3957
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3954..3956,3957..3959,3960..4673)
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3999..4020)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4260..4279)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4607..4628
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4796..4917
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4833..4852)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4887..4906
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4924..5379)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5061..5082)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5273..5292
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5406..5510
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5512..5869
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5512..5703
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5535..5555)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5720..5855
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5904..6698
                     /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             5904..6698
                     /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(5958..5977)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6568..6587
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6886..6905)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6930..6977
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7306..7894
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7795..7814
                     /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 gctagcatgg
      601 cggcgctgag cggtggcggt ggtggcggcg cggagccggg ccaggctctg ttcaacgggg
      661 acatggagcc cgaggccggc gccggcgccg gcgccgcggc ctcttcggct gcggaccctg
      721 ccattccgga ggaggtgtgg aatatcaaac aaatgattaa gttgacacag gaacatatag
      781 aggccctatt ggacaaattt ggtggggagc ataatccacc atcaatatat ctggaggcct
      841 atgaagaata caccagcaag ctagatgcac tccaacaaag agaacaacag ttattggaat
      901 ctctggggaa cggaactgat ttttctgttt ctagctctgc atcaatggat accgttacat
      961 cttcttcctc ttctagcctt tcagtgctac cttcatctct ttcagttttt caaaatccca
     1021 cagatgtggc acggagcaac cccaagtcac cacaaaaacc tatcgttaga gtcttcctgc
     1081 ccaacaaaca gaggacagtg gtacctgcaa ggtgtggagt tacagtccga gacagtctaa
     1141 agaaagcact gatgatgaga ggtctaatcc cagagtgctg tgctgtttac agaattcagg
     1201 atggagagaa gaaaccaatt ggttgggaca ctgatatttc ctggcttact ggagaagaat
     1261 tgcatgtgga agtgttggag aatgttccac ttacaacaca caactttgta cgaaaaacgt
     1321 ttttcacctt agcattttgt gacttttgtc gaaagctgct tttccagggt ttccgctgtc
     1381 aaacatgtgg ttataaattt caccagcgtt gtagtacaga agttccactg atgtgtgtta
     1441 attatgacca acttgatttg ctgtttgtct ccaagttctt tgaacaccac ccaataccac
     1501 aggaagaggc gtccttagca gagactgccc taacatctgg atcatcccct tccgcacccg
     1561 cctcggactc tattgggccc caaattctca ccagtccgtc tccttcaaaa tccattccaa
     1621 ttccacagcc cttccgacca gcagatgaag atcatcgaaa tcaatttggg caacgagacc
     1681 gatcctcatc agctcccaat gtgcatataa acacaataga acctgtcaat attgatgact
     1741 tgattagaga ccaaggattt cgtggtgatg gaggatcaac cacaggtttg tctgctaccc
     1801 cccctgcctc attacctggc tcactaacta acgtgaaagc cttacagaaa tctccaggac
     1861 ctcagcgaga aaggaagtca tcttcatcct cagaagacag gaatcgaatg aaaacacttg
     1921 gtagacggga ctcgagtgat gattgggaga ttcctgatgg gcagattaca gtgggacaaa
     1981 gaattggatc tggatcattt ggaacagtct acaagggaaa gtggggacca ggtggcagcg
     2041 gaggtcatac cttgtatgcg ccggggggtt atgacatcat gggttacctc atacagatca
     2101 tgaataggcc gaacccacaa gtggagctcg gacccgtcga tacctcctgc gctctcattc
     2161 tgtgtgacct taagcagaag gataccccta tcgtgtacgc ctccgaggca tttctgtaca
     2221 tgacagggta ctcgaacgcc gaagtgctgg gacggaactg ccgcttcctg caaagcccgg
     2281 atggaatggt gaagcctaag tcaacccgca aatacgtgga ctccaacact atcaacacca
     2341 tgcgcaaggc cattgaccgc aatgctgagg tgcaagtgga agtggtgaac ttcaagaaga
     2401 atggacagcg cttcgtcaac ttcctgacta tgatccccgt gcgcgacgag accggcgaat
     2461 accggtacag catggggttt cagtgtgaga cagagggcgg ctccggaggc agcggcggtt
     2521 ctggaggttc cggtggcggc tccggaggta gcggagggtc tcacactctt tacgcccctg
     2581 gaggatacga cattatggga tatttgattc agattatgaa ccgcccaaac cctcaggtcg
     2641 aactggggcc tgtggacacg tcatgtgccc tgatcctgtg cgatctgaag caaaaggaca
     2701 ctccgattgt ctacgcctcg gaagccttct tgtatatgac cggatacagc aatgcagagg
     2761 tgctcggtag gaactgcaga ttcctgcagt cccccgacgg gatggtgaaa ccaaagtcga
     2821 ctcgcaaata tgtggactcg aacacgatca atacaatgcg gaaggccatc gaccggaacg
     2881 ccgaggtcca ggtggaggtg gtcaacttta agaagaacgg ccagcggttc gtgaactttc
     2941 tcaccatgat tccggtccgg gatgaaaccg gagagtacag atactccatg ggattccagt
     3001 gcgaaaccga agggtccgga ggtcccggag gtgatgtggc agtgaaaatg ttgaatgtga
     3061 cagcacctac acctcagcag ttacaagcct tcaaaaatga agtaggagta ctcaggaaaa
     3121 cacgacatgt gaatatccta ctcttcatgg gctattccac aaagccacaa ctggctattg
     3181 ttacccagtg gtgtgagggc tccagcttgt atcaccatct ccatatcatt gagaccaaat
     3241 ttgagatgat caaacttata gatattgcac gacagactgc acagggcatg gattacttac
     3301 acgccaagtc aatcatccac agagacctca agagtaataa tatatttctt catgaagacc
     3361 tcacagtaaa aataggtgat tttggtctag ctacagagaa atctcgatgg agtgggtccc
     3421 atcagtttga acagttgtct ggatccattt tgtggatggc accagaagtc atcagaatgc
     3481 aagataaaaa tccatacagc tttcagtcag atgtatatgc atttggaatt gttctgtatg
     3541 aattgatgac tggacagtta ccttattcaa acatcaacaa cagggaccag ataattttta
     3601 tggtgggacg aggatacctg tctccagatc tcagtaaggt acggagtaac tgtccaaaag
     3661 ccatgaagag attaatggca gagtgcctca aaaagaaaag agatgagaga ccactctttc
     3721 cccaaattct cgcctctatt gagctgctgg cccgctcatt gccaaaaatt caccgcagtg
     3781 catcagaacc ctccttgaat cgggctggtt tccaaacaga ggattttagt ctatatgctt
     3841 gtgcttctcc aaaaacaccc atccaggcag ggggatatgg tgcgtttcct gtccacaagc
     3901 ttcgaattct gcagtcgacg gtaccgcggg cccgggatcc accggtcgcc accatggtga
     3961 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
     4021 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
     4081 tgaccctgaa gttgatctgc accaccggca agctgcccgt gccctggccc accctcgtga
     4141 ccaccctcgg ctacggcctg cagtgcttcg cccgctaccc cgaccacatg aagcagcacg
     4201 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
     4261 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     4321 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
     4381 agtacaacta caacagccac aacgtctata tcaccgccga caagcagaag aacggcatca
     4441 aggcaaactt caagatccgc cacaacatcg aggacggcgg cgtgcagctc gccgaccact
     4501 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     4561 gctaccagtc caaactgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     4621 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag taaagcggcc
     4681 gcgactctag atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa
     4741 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt
     4801 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     4861 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaagg
     4921 cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     4981 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     5041 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     5101 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     5161 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     5221 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     5281 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     5341 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa
     5401 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     5461 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg
     5521 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     5581 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     5641 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     5701 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     5761 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     5821 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag
     5881 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     5941 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     6001 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     6061 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga
     6121 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     6181 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     6241 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     6301 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     6361 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     6421 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     6481 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     6541 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     6601 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     6661 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     6721 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     6781 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     6841 ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga
     6901 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg
     6961 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac
     7021 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc
     7081 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat
     7141 agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     7201 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     7261 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     7321 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     7381 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     7441 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     7501 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     7561 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     7621 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     7681 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     7741 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     7801 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     7861 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     7921 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     7981 tgtggataac cgtattaccg ccatgcat
//
