LOCUS       sequence-366172-        8798 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Baculovirus expression of human KIF1C with P176L mutation and 6xHis 
            and GFP tags..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8798)
  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 8798)
  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: pFastBac-M13-6His-KIF1C_P176L-GFP
FEATURES             Location/Qualifiers
     source          1..8798
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      135..589
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(222..241)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     431..452
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        615..719
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(720..788,789..1580)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(938..957)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1751..2339
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2240..2259
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2493..2510
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  2644..2868
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(2935..3468)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3638..3643,3644..3660,3661..3666))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     promoter        4018..4109
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     4034..4051
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             4174..4191
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             4213..4233
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     regulatory      6475..6484
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      7573..7582
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(7579..7581,7582..7584,7585..8298)
                     /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(7579..7581,7582..7584,7585..8295)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(7624..7645)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(7885..7904)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     8232..8253
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    8421..8555
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8458..8477)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8512..8531
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  8584..8749
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 tccatgtgac aaaccgtcat cttcggctac tttttctctg tcacagaatg aaaatttttc
       61 tgtcatctct tcgttattaa tgtttgtaat tgactgaata tcaacgctta tttgcagcct
      121 gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
      181 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
      241 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
      301 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
      361 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
      421 gttcttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
      481 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
      541 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt
      601 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
      661 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
      721 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
      781 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
      841 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
      901 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
      961 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     1021 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     1081 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     1141 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     1201 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     1261 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     1321 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     1381 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     1441 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     1501 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     1561 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     1621 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     1681 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     1741 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     1801 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     1861 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     1921 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     1981 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     2041 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     2101 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     2161 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     2221 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     2281 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     2341 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     2401 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     2461 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     2521 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatagacc
     2581 agccgcgtaa cctggcaaaa tcggttacgg ttgagtaata aatggatgcc ctgcgtaagc
     2641 gggtgtgggc ggacaataaa gtcttaaact gaacaaaata gatctaaact atgacaataa
     2701 agtcttaaac tagacagaat agttgtaaac tgaaatcagt ccagttatgc tgtgaaaaag
     2761 catactggac ttttgttatg gctaaagcaa actcttcatt ttctgaagtg caaattgccc
     2821 gtcgtattaa agaggggcgt ggccaagggc atggtaaaga ctatattcgc ggcgttgtga
     2881 caatttaccg aacaactccg cggccgggaa gccgatctcg gcttgaacga attgttaggt
     2941 ggcggtactt gggtcgatat caaagtgcat cacttcttcc cgtatgccca actttgtata
     3001 gagagccact gcgggatcgt caccgtaatc tgcttgcacg tagatcacat aagcaccaag
     3061 cgcgttggcc tcatgcttga ggagattgat gagcgcggtg gcaatgccct gcctccggtg
     3121 ctcgccggag actgcgagat catagatata gatctcacta cgcggctgct caaacttggg
     3181 cagaacgtaa gccgcgagag cgccaacaac cgcttcttgg tcgaaggcag caagcgcgat
     3241 gaatgtctta ctacggagca agttcccgag gtaatcggag tccggctgat gttgggagta
     3301 ggtggctacg tctccgaact cacgaccgaa aagatcaaga gcagcccgca tggatttgac
     3361 ttggtcaggg ccgagcctac atgtgcgaat gatgcccata cttgagccac ctaactttgt
     3421 tttagggcga ctgccctgct gcgtaacatc gttgctgctg cgtaacatca aacatcgacc
     3481 cacggcgtaa cgcgcttgct gcttggatgc ccgaggcata gactgtacaa aaaaacagtc
     3541 ataacaagcc atgaaaaccg ccactgcgcc gttaccaccg ctgcgttcgg tcaaggttct
     3601 ggaccagttg cgtgagcgca tacgctactt gcattacagt ttacgaaccg aacaggctta
     3661 tgtcaactgg gttcgtgcct tcatccgttt ccacggtgtg cgtcacccgg caaccttggg
     3721 cagcagcgaa gtcgaggcat ttctgtcctg gctggcgaac gagcgcaagg tttcggtctc
     3781 cacgcatcgt caggcattgg cggccttgct gttcttctac ggcaaggtgc tgtgcacgga
     3841 tctgccctgg cttcaggaga tcggaagacc tcggccgtcg cggcgcttgc cggtggtgct
     3901 gaccccggat gaagtggttc gcatcctcgg ttttctggaa ggcgagcatc gtttgttcgc
     3961 ccaggactct agctatagtt ctagtggttg gctacgtata ctccggaata ttaatagatc
     4021 atggagataa ttaaaatgat aaccatctcg caaataaata agtattttac tgttttcgta
     4081 acagttttgt aataaaaaaa cctataaata ttccggatta ttcatacccc caccatcggg
     4141 cgcggatctc ggtccgaaac catgtcgtac taccatcacc atcaccatca cgattacgat
     4201 atcccaacga ccgaaaacct gtattttcag ggcgccatgg cgcgcccgga tctggctggt
     4261 gcctcggtga aagtggcagt gagggttcgg ccctttaacg cccgtgagac cagccaggat
     4321 gccaagtgtg tggtcagcat gcagggcaac accacctcca tcatcaatcc taaacagagc
     4381 aaggatgccc ccaaaagctt cacctttgac tactcctact ggtcacacac ttcgacggag
     4441 gacccccagt ttgcatctca gcagcaagtg tatcgggaca tcggagaaga gatgctgctc
     4501 cacgcctttg aaggctacaa cgtgtgcatc tttgcctatg ggcagaccgg ggctgggaaa
     4561 tcctatacca tgatggggcg acaggagcca gggcagcagg gcatcgtgcc ccagctctgt
     4621 gaggacctct tctctcgcgt tagtgagaac cagagtgctc agctatccta ctctgtggag
     4681 gtgagctata tggagatcta ctgtgagcgg gtacgagacc tcttgaaccc caagagtcgg
     4741 ggttctctgc gggtccggga gcaccccatc ctgggcctgt acgtgcagga cctgtccaaa
     4801 ttggctgtga cctcctacgc agacattgct gacctcatgg actgtggaaa taaagcacgg
     4861 actgtggctg ccaccaacat gaatgagacc agcagccgtt cccatgccgt ctttaccatc
     4921 gtcttcacac agcgctgcca tgaccagctc acggggctgg actcggagaa ggtcagtaag
     4981 atcagtttgg tggaccttgc tgggagtgag cgagccgact cctcaggggc ccggggcatg
     5041 cgcctgaagg aaggagccaa catcaataag tccctgacta cactagggaa agtgatctcg
     5101 gcccttgcag atatgcaatc aaagaagcga aagtcggatt ttatccccta cagggactct
     5161 gtgctcacct ggctgctcaa ggaaaatttg ggggggaact cacgcacagc catgattgca
     5221 gccctgagcc ctgctgacat caattacgag gagactctca gcaccctcag gtatgctgac
     5281 cgcaccaagc aaatccgctg caatgccatc atcaacgagg accctaatgc ccggctgatt
     5341 agagagctgc aggaggaagt agcccggctg cgggaactgc tgatggctca gggactgtca
     5401 gcctctgctc tggaaggcct gaagacggaa gaagggagtg tcagaggcgc cctgccagct
     5461 gtgtcatctc ccccagctcc agtttcaccc tcatcaccca ccacacataa tggggagctg
     5521 gagccgtcat tctcccccaa cacggagtcc cagattgggc ctgaggaagc catggagagg
     5581 ctgcaggaga cagagaagat tatagctgag ctgaacgaga catgggagga gaagctacgc
     5641 aagacagaag ccctgaggat ggagagagaa gcattgctgg ctgagatggg ggtggccgtc
     5701 cgggaggatg ggggaactgt gggcgtcttc tctccaaaga agactcccca cctggtgaac
     5761 ctgaacgaag accctctgat gtctgagtgt ctgctctacc acatcaaaga tggcgtcacc
     5821 agggtcggcc aagtagatat ggacatcaag ctgaccggac agttcattcg ggagcaacac
     5881 tgtctgttcc ggagcatccc ccagccagat ggagaagtgg tggtcactct ggagccttgt
     5941 gaaggagctg agacatatgt gaatgggaag cttgtgacgg agccgctggt gctgaagtca
     6001 gggaatagga ttgtgatggg caagaaccac gttttccgct tcaaccaccc ggagcaggca
     6061 aggctggaac gggaacgagg ggtcccccca cccccaggac cgccctctga gccagtcgac
     6121 tggaactttg cccagaagga actgctggag cagcaaggca tcgacataaa gctggaaatg
     6181 gagaagaggc tgcaggatct ggagaatcag taccggaaag aaaaggaaga agccgatctt
     6241 ctgctggagc agcagcgact gtatgcagac tcggacagcg gggatgactc tgacaagcgc
     6301 tcttgtgaag agagctggag gctcatctcc tccttgcggg agcagctgcc gcccaccacg
     6361 gtccagacca ttgtcaaacg ctgtggtctg cccagcagtg gcaagcgcag ggcccctcgc
     6421 agggtttatc agatccccca gcgacgcagg ctgcagggca aagacccccg ctgggccacc
     6481 atggctgacc tgaagatgca ggcggtgaag gagatctgct acgaggtggc cctggctgac
     6541 ttccgccacg ggcgggctga gattgaggcc ctggccgccc tcaagatgcg ggagctgtgt
     6601 cgcacctatg gcaagccaga cggccccgga gacgcctgga gggctgtggc ccgggatgtc
     6661 tgggacactg taggcgagga ggaaggaggt ggagctggca gtggtggtgg cagtgaggag
     6721 ggagcccgag gggcggaggt ggaggacctc cgggcccaca tcgacaagct gacggggatt
     6781 ctgcaggagg tgaagctgca gaacagcagc aaggaccggg agctgcaggc cctgcgggac
     6841 cgcatgctcc gcatggagag ggtcatcccc ctggcccagg atcatgagga tgagaatgaa
     6901 gaaggtggtg aggtcccctg ggccccgcct gaaggatcag aggcagcaga ggaggcagcc
     6961 cccagtgacc gcatgccgtc agcccggccc ccctcgccac cactgtcaag ctgggagcgg
     7021 gtgtcacggc tcatggagga ggaccctgcc ttccgtcgtg gtcgtcttcg ctggctcaag
     7081 caggagcagc tacggctgca gggactgcag ggctctgggg gccggggcgg ggggctgcgc
     7141 aggcccccag cccgctttgt gccccctcac gactgcaagc tacgcttccc cttcaagagc
     7201 aacccccagc accgggagtc ttggccaggg atggggagcg gggaggctcc aactccgctc
     7261 caaccccctg aggaggtcac tccccatcca gccacccctg cccgccggcc tccgagtccc
     7321 cgaaggtccc accatccccg caggaactcc ctggatggag ggggccgatc ccggggagcg
     7381 ggttctgcac agcctgaacc ccagcacttc cagcccaaaa agcacaactc ttatccccag
     7441 ccaccccaac cctacccagc ccagcggccc ccagggcccc gctacccccc atacactact
     7501 cccccacgaa tgagacggca gcgttctgcc cctgacctca aggagagtgg ggcagctgtg
     7561 gatccaccgg tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
     7621 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc
     7681 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg
     7741 cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc
     7801 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
     7861 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag
     7921 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac
     7981 ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg
     8041 gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac
     8101 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
     8161 ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag
     8221 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg
     8281 gacgagctgt acaagtaaag cggccgcgac tctagatcat aatcagccat accacatttg
     8341 tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa
     8401 tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca
     8461 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     8521 ccaaactcat caatgtatct tatcatgtct ggatctgatc actgatatcg cctaggagat
     8581 ccgaaccaga taagtgaaat ctagttccaa actattttgt catttttaat tttcgtatta
     8641 gcttacgacg ctacacccag ttcccatcta ttttgtcact cttccctaaa taatccttaa
     8701 aaactccatt tccacccctc ccagttccca actattttgt ccgcccacag cggggcattt
     8761 ttcttcctgt tatgttttta atcaaacatc ctgccaac
//
