LOCUS       sequence-198855-        6796 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses fusion of kinesin heavy chain 5A (1-572) with GFP and CIB1
            (1-170).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6796)
  AUTHORS   Duan L, Che D, Zhang K, Ong Q, Guo S, Cui B
  TITLE     Optogenetic control of molecular motors and organelle distributions 
            in cells.
  JOURNAL   Chem Biol. 2015 May 21;22(5):671-82. doi: 
            10.1016/j.chembiol.2015.04.014. Epub 2015 May 9.
  PUBMED    25963241
REFERENCE   2  (bases 1 to 6796)
  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"; doi: 
            "10.1016/j.chembiol.2015.04.014"; journalName: "Chem Biol"; date: 
            "2015-05-21- 21"; volume: "22"; issue: "5"; pages: "671-82"
COMMENT     Sequence Label: Kif5A-GFP-CIBN
FEATURES             Location/Qualifiers
     source          1..6796
                     /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      606..615
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2334..2336,2337..2339,2340..3050)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2334..2336,2337..2339,2340..3050)
                     /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(2379..2400)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2640..2659)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2987..3008
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3057..3566
                     /codon_start=1
                     /product="N-terminal portion of the CIB1 transcription 
                     factor from Arabidopsis thaliana"
                     /label=CIBN
                     /note="binds to CRY2 (cryptochrome 2) that has absorbed 
                     blue light (Kennedy et al., 2010)"
                     /translation="MNGAIGGDLLLNFPDMSVLERQRAHLKYLNPTFDSPLAGFFADSS
                     MITGGEMDSYLSTAGLNLPMMYGETTVEGDSRLSISPETTLGTGNFKAAKFDTETKDCN
                     EAAKKMTMNRDDLVEEGEEEKSKITEQNNGSTKSIKKMKHKAKKEENNFSNDSSKVTKE
                     LEKTDYI"
     polyA_signal    3584..3705
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3621..3640)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3675..3694
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3712..4167)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3849..3870)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4061..4080
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4194..4298
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4300..4657
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4300..4491
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4323..4343)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4508..4643
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4692..5486
                     /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             4692..5486
                     /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(4746..4765)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5356..5375
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5674..5693)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5718..5765
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6094..6682
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6583..6602
                     /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 gctagcgcta
      601 agcttgccac catgggcgcg cctgaattcg cggagaccaa taacgaatgc agcatcaagg
      661 tgctttgccg atttcggccc ctgaaccagg ccgagatcct gcggggggac aagttcatcc
      721 ccattttcca aggggacgac agcgtcatta ttgggggaaa gccatatgtc ttcgaccgtg
      781 tattcccccc aaacacgact caggagcagg tttaccacgc ctgtgccatg cagatcgtca
      841 aagacgtcct tgctggctac aacggcacaa tttttgctta tggacagaca tcctcaggga
      901 aaacgcacac catggaggga aagctgcacg accctcagct catgggcatc attccccgga
      961 tcgcccgaga catcttcaac cacatctact ccatggatga gaaccttgag ttccacatta
     1021 aggtgtctta ctttgagatt tacctggata agatccgtga ccttttggat gtgaccaaga
     1081 caaacctgtc tgtgcatgag gacaaaaacc gggtgccgtt tgtcaagggt tgtaccgaac
     1141 gctttgtgtc cagcccagag gagattctgg atgtgattga tgaggggaag tcgaaccgtc
     1201 acgttgccgt caccaacatg aatgagcaca gctctcggag tcacagcatc ttcctcatca
     1261 acatcaagca ggagaacata gagacggagc agaagctcag cgggaaactg taccttgtgg
     1321 atctggccgg aagcgagaag gtcagcaaga caggagcaga gggagccgtt ctggatgagg
     1381 caaagaatat caataagtcg ctgtcggccc tggggaatgt gatctctgct ctggccgagg
     1441 gcactaaaag ctacgtgccg taccgtgaca gcaaaatgac gaggattctc caggactctc
     1501 tgggagggaa ctgcaggacc accatgttca tctgctgctc accatccagc tacaatgacg
     1561 cagagaccaa gtccacactc atgtttgggc agcgggcaaa gaccatcaag aacactgcct
     1621 cagtgaatct ggagttgact gctgagcaat ggaagaagaa atatgagaag gagaaggaga
     1681 agaccaaggc ccagaaggag acaattgcga agctggaggc tgagcttagc cggtggcgca
     1741 atggagagaa tgtgcctgag actgagcgcc tggctggaga ggactcagct ctggcagccg
     1801 agatctgtga ggagacccct gtgaatgaca actcatccat cgtggtacgc atcgcaccgg
     1861 aggaaaggca gaaatacgag gaagagatcc gccgtctcta caagcagctc gacgacaagg
     1921 atgatgagat caaccagcag agccaactca ttgagaagct gaagcagcag atgctggacc
     1981 aggaagagct gctagtgtcc actcggggag acaatgagaa ggtccagcgg gagctgagcc
     2041 acctgcagtc tgagaacgat gccgcgaagg aggaggtgaa ggaagtgttg caggccctgg
     2101 aggagctggc agtcaactat gaccagaagt cccaggaggt ggaggagaag agccagcaga
     2161 accagctgct ggtggatgag ctgtcccaga aagtggccac catgctgtcc ctggagtctg
     2221 agctacagcg gctccaggaa gtcagtggac accagcgaaa acggattgct gaggtgctga
     2281 atggactgat gaaggacctg agtgagttca gtgtcatcgt gggcaagtcg actatggtga
     2341 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
     2401 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
     2461 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga
     2521 ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg
     2581 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
     2641 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     2701 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
     2761 agtacaacta caacagccac aacgtctata tcatggccga caagcagaag aacggcatca
     2821 aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact
     2881 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     2941 gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     3001 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag ggatccatga
     3061 atggagctat aggaggtgac cttttgctca attttcctga catgtcggtc ctagagcgcc
     3121 aaagggctca cctcaagtac ctcaatccca cctttgattc tcctctcgcc ggcttctttg
     3181 ccgattcttc aatgattacc ggcggcgaga tggacagcta tctttcgact gccggtttga
     3241 atcttccgat gatgtacggt gagacgacgg tggaaggtga ttcaagactc tcaatttcgc
     3301 cggaaacgac gcttgggact ggaaatttca aggcagcgaa gtttgataca gagactaagg
     3361 attgtaatga ggcggcgaag aagatgacga tgaacagaga tgacctagta gaagaaggag
     3421 aagaagagaa gtcgaaaata acagagcaaa acaatgggag cacaaaaagc atcaagaaga
     3481 tgaaacacaa agccaagaaa gaagagaaca atttctctaa tgattcatct aaagtgacga
     3541 aggaattgga gaaaacggat tatatttaac aattgttgtt gttaacttgt ttattgcagc
     3601 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     3661 actgcattct agttgtggtt tgtccaaact catcaatgta tcttaaggcg taaattgtaa
     3721 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     3781 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     3841 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     3901 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     3961 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     4021 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     4081 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     4141 cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa
     4201 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4261 cctgataaat gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct
     4321 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     4381 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     4441 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     4501 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     4561 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     4621 gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag
     4681 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     4741 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     4801 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     4861 tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     4921 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     4981 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     5041 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     5101 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     5161 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga
     5221 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     5281 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     5341 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     5401 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     5461 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     5521 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     5581 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     5641 ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga
     5701 aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt
     5761 tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc
     5821 ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg
     5881 tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta
     5941 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     6001 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     6061 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     6121 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     6181 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     6241 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     6301 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     6361 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     6421 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     6481 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     6541 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     6601 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     6661 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     6721 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     6781 tattaccgcc atgcat
//
