LOCUS       sequence-366169-        8023 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human KIF1C with P176L mutation and GFP 
            tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8023)
  AUTHORS   Siddiqui N, Roth D, Toleikis A, Zwetsloot AJ, Cross RA, Straube A
  TITLE     Force generation of KIF1C is impaired by pathogenic mutations.
  JOURNAL   Curr Biol. 2022 Sep 12;32(17):3862-3870.e6. doi: 
            10.1016/j.cub.2022.07.029. Epub 2022 Aug 11.
  PUBMED    35961316
REFERENCE   2  (bases 1 to 8023)
  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.cub.2022.07.029"; journalName: "Curr Biol"; date: 
            "2022-09-12- 12"; volume: "32"; issue: "17"; pages: "3862-3870.e6"
COMMENT     Sequence Label: pKIF1C_P176L-GFP
FEATURES             Location/Qualifiers
     source          1..8023
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             74..868
                     /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             74..868
                     /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(128..147)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     738..757
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1056..1075)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1100..1147
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1476..2064
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1965..1984
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2239..2542
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2543..2746
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2697..2717
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      5043..5052
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      6141..6150
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(6147..6149,6150..6152,6153..6866)
                     /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(6147..6149,6150..6152,6153..6863)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(6192..6213)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(6453..6472)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6800..6821
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    6989..7110
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7026..7045)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7080..7099
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(7117..7572)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7254..7275)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7466..7485
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        7599..7703
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(7705..8023,1..39)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        7705..7896
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(7728..7748)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      join(7913..8023,1..25)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag agacaggatg
       61 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
      121 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
      181 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
      241 cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
      301 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
      361 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
      421 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
      481 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
      541 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
      601 gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
      661 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
      721 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
      781 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
      841 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
      901 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
      961 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     1021 ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga gacaataccg
     1081 gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg tgttgggtcg
     1141 tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga
     1201 ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg
     1261 ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat agcctcaggt
     1321 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     1381 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     1441 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     1501 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     1561 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     1621 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     1681 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     1741 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     1801 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     1861 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     1921 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     1981 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     2041 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     2101 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     2161 cgtattaccg ccatgcatta gttattaata gtaatcaatt acggggtcat tagttcatag
     2221 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     2281 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     2341 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     2401 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
     2461 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
     2521 attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata
     2581 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
     2641 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
     2701 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctggtt tagtgaaccg
     2761 tcagatccgc tagcgctacc ggactcagat ctcgagctca agcttcgaat tctggagcta
     2821 tggctggtgc ctcggtgaaa gtggcagtga gggttcggcc ctttaacgcc cgtgagacca
     2881 gccaggatgc caagtgtgtg gtcagcatgc agggcaacac cacctccatc atcaatccta
     2941 aacagagcaa ggatgccccc aaaagcttca cctttgacta ctcctactgg tcacacactt
     3001 cgacggagga cccccagttt gcatctcagc agcaagtgta tcgggacatc ggagaagaga
     3061 tgctgctcca cgcctttgaa ggctacaacg tgtgcatctt tgcctatggg cagaccgggg
     3121 ctgggaaatc ctataccatg atggggcgac aggagccagg gcagcagggc atcgtgcccc
     3181 agctctgtga ggacctcttc tctcgcgtta gtgagaacca gagtgctcag ctatcctact
     3241 ctgtggaggt gtcttacatg gaaatctact gtgagcgggt acgagacctc ttgaacccca
     3301 agagtcgggg ttctctgcgg gtccgggagc accccatcct gggcctgtac gtgcaggacc
     3361 tgtccaaatt ggctgtgacc tcctacgcag acattgctga cctcatggac tgtggaaata
     3421 aagcacggac tgtggctgcc accaacatga atgagaccag cagccgttcc catgccgtct
     3481 ttaccatcgt cttcacacag cgctgccatg accagctcac ggggctggac tcggagaagg
     3541 tcagtaagat cagtttggtg gaccttgctg ggagtgagcg agccgactcc tcaggggccc
     3601 ggggcatgcg cctgaaggaa ggagccaaca tcaataagtc cctgactaca ctagggaaag
     3661 tgatctcggc ccttgcagat atgcaatcaa agaagcgaaa gtcggatttt atcccctaca
     3721 gggactctgt gctcacctgg ctgctcaagg aaaatttggg ggggaactca cgcacagcca
     3781 tgattgcagc cctgagccct gctgacatca attacgagga gactctcagc accctcaggt
     3841 atgctgaccg caccaagcaa atccgctgca atgccatcat caacgaggac cctaatgccc
     3901 ggctgattag agagctgcag gaggaagtag cccggctgcg ggaactgctg atggctcagg
     3961 gactgtcagc ctctgctctg gaaggcctga agacggaaga agggagtgtc agaggcgccc
     4021 tgccagctgt gtcatctccc ccagctccag tttcaccctc atcacccacc acacataatg
     4081 gggagctgga gccgtcattc tcccccaaca cggagtccca gattgggcct gaggaagcca
     4141 tggagaggct gcaggagaca gagaagatta tagctgagct gaacgagaca tgggaggaga
     4201 agctacgcaa gacagaagcc ctgaggatgg agagagaagc attgctggct gagatggggg
     4261 tggccgtccg ggaggatggg ggaactgtgg gcgtcttctc tccaaagaag actccccacc
     4321 tggtgaacct gaacgaagac cctctgatgt ctgagtgtct gctctaccac atcaaagatg
     4381 gcgtcaccag ggtcggccaa gtagatatgg acatcaagct gaccggacag ttcattcggg
     4441 agcaacactg tctgttccgg agcatccccc agccagatgg agaagtggtg gtcactctgg
     4501 agccttgtga aggagctgag acatatgtga atgggaagct tgtgacggag ccgctggtgc
     4561 tgaagtcagg gaataggatt gtgatgggca agaaccacgt tttccgcttc aaccacccgg
     4621 agcaggcaag gctggaacgg gaacgagggg tccccccacc cccaggaccg ccctctgagc
     4681 cagtcgactg gaactttgcc cagaaggaac tgctggagca gcaaggcatc gacataaagc
     4741 tggaaatgga gaagaggctg caggatctgg agaatcagta ccggaaagaa aaggaagaag
     4801 ccgatcttct gctggagcag cagcgactgt atgcagactc ggacagcggg gatgactctg
     4861 acaagcgctc ttgtgaagag agctggaggc tcatctcctc cttgcgggag cagctgccgc
     4921 ccaccacggt ccagaccatt gtcaaacgct gtggtctgcc cagcagtggc aagcgcaggg
     4981 cccctcgcag ggtttatcag atcccccagc gacgcaggct gcagggcaaa gacccccgct
     5041 gggccaccat ggctgacctg aagatgcagg cggtgaagga gatctgctac gaggtggccc
     5101 tggctgactt ccgccacggg cgggctgaga ttgaggccct ggccgccctc aagatgcggg
     5161 agctgtgtcg cacctatggc aagccagacg gccccggaga cgcctggagg gctgtggccc
     5221 gggatgtctg ggacactgta ggcgaggagg aaggaggtgg agctggcagt ggtggtggca
     5281 gtgaggaggg agcccgaggg gcggaggtgg aggacctccg ggcccacatc gacaagctga
     5341 cggggattct gcaggaggtg aagctgcaga acagcagcaa ggaccgggag ctgcaggccc
     5401 tgcgggaccg catgctccgc atggagaggg tcatccccct ggcccaggat catgaggatg
     5461 agaatgaaga aggtggtgag gtcccctggg ccccgcctga aggatcagag gcagcagagg
     5521 aggcagcccc cagtgaccgc atgccgtcag cccggccccc ctcgccacca ctgtcaagct
     5581 gggagcgggt gtcacggctc atggaggagg accctgcctt ccgtcgtggt cgtcttcgct
     5641 ggctcaagca ggagcagcta cggctgcagg gactgcaggg ctctgggggc cggggcgggg
     5701 ggctgcgcag gcccccagcc cgctttgtgc cccctcacga ctgcaagcta cgcttcccct
     5761 tcaagagcaa cccccagcac cgggagtctt ggccagggat ggggagcggg gaggctccaa
     5821 ctccgctcca accccctgag gaggtcactc cccatccagc cacccctgcc cgccggcctc
     5881 cgagtccccg aaggtcccac catccccgca ggaactccct ggatggaggg ggccgatccc
     5941 ggggagcggg ttctgcacag cctgaacccc agcacttcca gcccaaaaag cacaactctt
     6001 atccccagcc accccaaccc tacccagccc agcggccccc agggccccgc taccccccat
     6061 acactactcc cccacgaatg agacggcagc gttctgcccc tgacctcaag gagagtgggg
     6121 cagctgtgga tccaccggtc gccaccatgg tgagcaaggg cgaggagctg ttcaccgggg
     6181 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg
     6241 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     6301 gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct
     6361 tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     6421 gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg
     6481 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca
     6541 aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct
     6601 atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca
     6661 tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg
     6721 gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc
     6781 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc
     6841 tcggcatgga cgagctgtac aagtaaagcg gccgcgactc tagatcataa tcagccatac
     6901 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     6961 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     7021 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     7081 tggtttgtcc aaactcatca atgtatctta aggcgtaaat tgtaagcgtt aatattttgt
     7141 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     7201 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     7261 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     7321 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     7381 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     7441 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     7501 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     7561 tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     7621 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     7681 aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga atgtgtgtca
     7741 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     7801 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     7861 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     7921 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     7981 tgcagaggcc gaggccgcct cggcctctga gctattccag aag
//
