LOCUS       sequence-435689-        7368 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of CLIPf-KIF5A(411-680)-SNAPf-HisTag-FLAG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7368)
  AUTHORS   Smith ER, Turner ED, Abdelhamid MAS, Craggs TD, Twelvetrees AE
  TITLE     Kinesin-1 is highly flexible and adopts an open conformation in the 
            absence of cargo
  JOURNAL   bioRxiv 2024.12.20.629623
REFERENCE   2  (bases 1 to 7368)
  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: "bioRxiv 
            2024.12.20.629623"
COMMENT     Sequence Label: CLIPf-KIF5A(411-680)-SNAPf-HisTag-FLAG
FEATURES             Location/Qualifiers
     source          1..7368
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    89..210
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(126..145)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     180..199
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(259..275)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(259..275)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(272..294)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    283..299
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(307..313,314..331,332..337))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    352..373
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(490..507)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(661..1249)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(741..760)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1420..2211,2212..2280))
                     /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     2043..2062
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2281..2385)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2460..2479)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2651..3030
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3031..3234
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3185..3205
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3279..3298
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3279..3297
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5286..5303
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5304..5327
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(5375..5392)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5381..5605
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5651..6079
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5738..5757)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5948..5969
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6088..6108)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6093..6422
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6273..6408
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6335..6354
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             6489..7283
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6543..6562)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7153..7172
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
       61 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
      121 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
      181 tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta
      241 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
      301 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
      361 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
      421 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
      481 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
      541 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
      601 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
      661 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
      721 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
      781 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
      841 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
      901 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
      961 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     1021 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     1081 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     1141 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     1201 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     1261 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     1321 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     1381 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     1441 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     1501 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     1561 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     1621 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     1681 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     1741 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     1801 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     1861 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     1921 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     1981 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     2041 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     2101 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     2161 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     2221 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     2281 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     2341 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     2401 agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt
     2461 acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag
     2521 gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat
     2581 tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga
     2641 tatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
     2701 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
     2761 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     2821 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     2881 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     2941 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     3001 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     3061 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     3121 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     3181 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg
     3241 ctaactagag aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag
     3301 acccaagctg gctagcgttt aaacttaagc ttggtaccat ggacaaggac tgcgagatga
     3361 agcggaccac actggatagc cctctgggca agctggaact gagcggctgt gaacagggcc
     3421 tgcaccggat catctttctc ggcaagggaa cctctgccgc cgatgctgtt gaagtgcctg
     3481 ctcctgctgc tgtgctcgga ggacctgaac ctctgattca ggccaccgcc tggctgaacg
     3541 cctactttca tcagcccgag gccatcgagg agttccccgt tcctgctctg catcaccccg
     3601 tgttccagca agaaagcttc accagacagg tgctgtggaa gctgctgaag gtggtcaagt
     3661 tcggcgaagt gatcagcgag tctcacctgg ccgctctcgt gggaaatcca gctgctacag
     3721 ccgccgtgaa cacagccctg gatggaaacc ctgtgcctat tctgatcccc tgccacagag
     3781 tggtgcaggg cgattctgat gtgggccctt atcttggagg cctggccgtg aaagaatggc
     3841 tgctggccca cgaaggacac agactgggaa aacctggcct cggaggatcc cctgaggaaa
     3901 ggcagaaata tgaggaagag atccgccgtc tctacaagca gcttgatgac aaggatgatg
     3961 agatcaacca gcagagccag ctcattgaga agctgaagca gcagatgctg gaccaggaag
     4021 agctgctcgt gtccactcgg ggagacaacg agaaggtcca gcgggagctt agccacctgc
     4081 agtccgagaa cgatgctgcg aaggacgagg tgaaggaagt gctgcaggcc ctagaggagc
     4141 tggcggtcaa ctacgaccag aagtcccagg aggtggagga gaagagccag cagaaccagc
     4201 tgctggtgga cgagctgtcc cagaaagtgg ccaccatgct gtccctggag tccgagctac
     4261 agcggctcca ggaggtcagt ggacaccagc gaaagcggat cgctgaggtg ctgaatgggc
     4321 tgatgaggga cctgagtgag ttcagtgtca tcgtgggcaa cggcgagatt aagctgcccg
     4381 tggagatcag tggggccatc gaggaggagt tcacggtggc ccggctctac atcagcaaga
     4441 tcaagtcgga ggtgaagtcc gtggttaagc gatgtcggca gctggagaac ctccaggtgg
     4501 agtgtcatcg caagatggag gtgaccggta gggagctgtc atcttgccaa ctgctcatct
     4561 cacagcatga ggccaagatc cgttcactca cggagtacat gcagactgtg gagttgaaga
     4621 aacggcacct ggaagagtcc tacgactccc tgagcgatga gcttgccagg ctccaggcgc
     4681 acgaaactgt acacgaggta gaattcgata aggattgtga aatgaagcgc acaaccctgg
     4741 actccccact gggcaaactt gagctgtctg gatgcgagca gggactgcac agaatcatat
     4801 tcctcggaaa gggcaccagc gccgctgatg cagtggaagt tccagcacca gctgcagtgc
     4861 ttggtggacc agagccactt atgcaggcta cagcttggct gaatgcttat ttccaccagc
     4921 cggaagccat cgaagagttt cccgtgccag cactgcacca tcctgtcttt caacaagagt
     4981 ccttcacacg gcaagtcctg tggaaactcc tgaaagtcgt gaaatttggg gaagtgatct
     5041 cctacagcca cctggctgca ctggccggaa atcctgcagc aactgccgct gtgaaaactg
     5101 ccctgtccgg caatccagtg ccaatcctga ttccttgtca ccgggtcgtc cagggcgacc
     5161 tggatgttgg aggatatgaa ggcggactgg ctgtcaaaga gtggctcctt gctcacgagg
     5221 gccatagact cggaaagcct ggacttggtg gaagcgccgg atctgcagca ggatctggcg
     5281 agtttcacca ccaccatcac cacgactaca aggacgacga cgacaaatga gcggccgctc
     5341 gagtctagag ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag
     5401 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
     5461 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
     5521 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat
     5581 gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg
     5641 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     5701 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     5761 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     5821 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     5881 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     5941 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     6001 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     6061 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     6121 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     6181 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     6241 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     6301 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     6361 ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     6421 aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat
     6481 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     6541 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     6601 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     6661 atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     6721 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     6781 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     6841 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     6901 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     6961 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca
     7021 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     7081 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     7141 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     7201 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     7261 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     7321 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatga
//
