LOCUS       sequence-346983-        8330 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Non-processive monomeric KIF1A for optogenetic heterodimerization to
            iLID via SSPB(micro).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8330)
  AUTHORS   Nijenhuis W, van Grinsven MMP, Kapitein LC
  TITLE     An optimized toolbox for the optogenetic control of intracellular 
            transport.
  JOURNAL   J Cell Biol. 2020 Apr 6;219(4). pii: 133834. doi: 
            10.1083/jcb.201907149.
  PUBMED    32328628
REFERENCE   2  (bases 1 to 8330)
  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: "J Cell 
            Biol."; date: "2020-04-6"; volume: "219(4). pii: 133834. doi"; 
            pages: " 10.1083/jcb.201907149"
COMMENT     Sequence Label: pB80-KIF1A(1-365)-GFP-SSPB(micro)
FEATURES             Location/Qualifiers
     source          1..8330
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1064..1086
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     regulatory      1149..1158
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2131..2140
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2349..2351,2352..3062)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2349..2351,2352..3062)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2391..2412)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2652..2671)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2999..3020
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             join(3192..3398,3399..3401,3402..3521)
                     /codon_start=1
                     /product="SspB from Haemophilus influenzae, with the 
                     mutations Y11K and A15E to disrupt the natural dimerization
                     of the protein and with the mutation R74Q to reduce the 
                     affinity for iLID"
                     /label=SspB micro
                     /note="dimerizes with iLID at moderate affinity to create 
                     iLID micro (Guntas et al., 2015)"
                     /translation="SSPKRPKLLREYYDWLVDNSFTPYLVVDATYLGVNVPVEYVKDGQ
                     IVLNLSASATGNLQLTNDFIQFNAQFKGVSRELYIPMGAALAIYARENGDGVMFEPEEI
                     YDELNI"
     polyA_signal    3654..3788
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3691..3710)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3745..3764
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3802..3818)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3802..3818)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3815..3837)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3826..3842
                     /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(3850..3856,3857..3874,3875..3880))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3895..3916
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4033..4050)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4204..4792)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4284..4303)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4963..5754,5755..5823))
                     /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     5586..5605
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5824..5928)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5996..6014
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6052..6074)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6174..6193
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6387..6409
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6401..6418
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6402..6418
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
       61 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      121 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      181 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      241 tccgggctgt aattagcgct tggtttaatg acggctcgtt tcttttctgt ggctgcgtga
      301 aagccttaaa gggctccggg agggcccttt gtgcgggggg gagcggctcg gggggtgcgt
      361 gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggctccgc gctgcccggc ggctgtgagc
      421 gctgcgggcg cggcgcgggg ctttgtgcgc tccgcagtgt gcgcgagggg agcgcggccg
      481 ggggcggtgc cccgcggtgc ggggggggct gcgaggggaa caaaggctgc gtgcggggtg
      541 tgtgcgtggg ggggtgagca gggggtgtgg gcgcggcggt cgggctgtaa cccccccctg
      601 cacccccctc cccgagttgc tgagcacggc ccggcttcgg gtgcggggct ccgtgcgggg
      661 cgtggcgcgg ggctcgccgt gccgggcggg gggtggcggc aggtgggggt gccgggcggg
      721 gcggggccgc ctcgggccgg ggagggctcg ggggaggggc gcggcggccc ccggagcgcc
      781 ggcggctgtc gaggcgcggc gagccgcagc cattgccttt tatggtaatc gtgcgagagg
      841 gcgcagggac ttcctttgtc ccaaatctgt gcggagccga aatctgggag gcgccgccgc
      901 accccctcta gcgggcgcgg ggcgaagcgg tgcggcgccg gcaggaagga aatgggcggg
      961 gagggccttc gtgcgtcgcc gcgccgccgt ccccttctcc atctccagcc tcggggctgt
     1021 ccgcaggggg acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt
     1081 gtgaccggcg gggtttatat cttcccttct ctgttcctcc gcagccccca agctaggcaa
     1141 ttgggcgcgc caccatggcc ggggcttcgg tgaaggtggc ggtgcgtgtc cgccccttca
     1201 actcccggga aatgagccga gactccaagt gcatcattca gatgtctgga agcaccacca
     1261 ctattgtcaa ccccaaacag cccaaggaga cacccaagag cttcagcttt gattactcct
     1321 actggtcaca tacctcgccg gaggacatca actatgcctc tcagaagcag gtctacaggg
     1381 acattgggga agagatgcta cagcatgcct ttgagggcta caatgtgtgc atctttgcct
     1441 atgggcagac gggggccggc aagtcctaca ccatgatggg gaaacaggag aaggaccagc
     1501 aaggcatcat tccacagctc tgcgaagacc tcttctctcg aatcaacgac accaccaacg
     1561 acaacatgtc ctactctgtg gaggtcagct acatggagat ttactgtgag cgagtccgtg
     1621 acctcctgaa ccccaagaac aaaggcaacc tgcgtgtgag ggagcaccct ctgctggggc
     1681 cttatgtgga ggacctctcc aagctggctg taacctccta caatgacatc caggacctca
     1741 tggactcagg gaacaaggcc aggactgtgg cggccacaaa catgaatgag accagcagcc
     1801 gttctcacgc agtcttcaac atcatcttta cccagaagcg ccatgatgca gagaccaaca
     1861 tcaccacaga gaaagtgagc aaaatcagcc tggtggacct ggcaggcagt gaacgagccg
     1921 actccacagg agccaagggc acgcgactca aggagggggc caacatcaac aagtctttga
     1981 ccaccctggg aaaggtcatc tctgctctgg ctgaaatgga ctctggaccc aacaagaaca
     2041 agaaaaagaa gaagacagat ttcatccctt accgagactc cgtattgaca tggcttctcc
     2101 gggaaaacct cggtggcaac tcaaggacag ccatggtggc agccctgagc cctgcggaca
     2161 tcaactatga tgagaccctc agcacactga gatatgcaga tcgggccaaa caaatccgct
     2221 gcaatgccat catcaatgaa gaccccaact ctagaggtgg cgggggcagt gggggagggg
     2281 gaagtggagg cggcggttct ggcgggggag gctccggtgg cggggggagc gggggtggga
     2341 gtaagcttgt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2401 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     2461 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     2521 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     2581 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     2641 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     2701 ccctggtgaa ccgcattgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     2761 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     2821 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     2881 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     2941 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     3001 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     3061 aggctagcgg aggcggtggc tcaggcggag gagggtcagg agggggaggc agtggcgggg
     3121 gcggctctgg aggagggggt tctggtggtg gtagctacgt agatctgagc ggcctgaccc
     3181 tgcaggaatt cagctccccg aaacgcccta agctgctgcg tgaatattac gattggctgg
     3241 ttgataacag ctttacccca tatctggtgg tggatgccac atacctgggc gtgaacgtgc
     3301 ccgtggagta tgtgaaagac ggtcagatcg tgctgaatct gtctgcaagt gcgaccggca
     3361 acctgcaact gacaaatgat tttatccagt tcaacgccca gtttaagggc gtgtctcgtg
     3421 aactgtatat cccgatgggt gccgctctgg ccatttacgc tcgcgagaac ggcgatggtg
     3481 tgatgttcga accagaagaa atctatgacg agctgaatat tggttaaacc ggtcctagat
     3541 aagtaatgat cataatcagc catatcacat ctgtagaggt tttacttgct ttaaaaaacc
     3601 tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt
     3661 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     3721 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     3781 tctggatcaa ttcgtaatca tgtcatagct gtttcctgtg tgaaattgtt atccgctcac
     3841 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     3901 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     3961 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     4021 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     4081 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     4141 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4201 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4261 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4321 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4381 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4441 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4501 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4561 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4621 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     4681 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     4741 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     4801 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     4861 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     4921 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     4981 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     5041 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     5101 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     5161 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     5221 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5281 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5341 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5401 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     5461 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     5521 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     5581 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     5641 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     5701 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     5761 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     5821 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     5881 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     5941 aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag
     6001 gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     6061 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     6121 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc
     6181 agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag
     6241 gagaaaatac cgcatcaggc gccattcgcc attcaggctg cgcaactgtt gggaagggcg
     6301 atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg
     6361 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtgc
     6421 caagttggga tctttgcatt ggcccacggc tctcaggatg gggatgctcc ccttcagcac
     6481 ccggttcccc ttggaaactg atggtcctgg ctctgtggca tggcagtggc actgtgagga
     6541 gccccctacc agcagcacac agtggggttg gcactgccac gctccggatg ccgcgctctg
     6601 atccaacccc ataatcaagg gaacccgaat tgccccatca ttgcccccac cacccccatc
     6661 ctgccgggcc ctcacacccc acgctgcctt ctggtgacat tccccagccc aaacccacgg
     6721 cttcatggct accgcggggc atttcccatt gccgccccat tatcagctct gcacacctcc
     6781 cgctgtaccc atgcctcgtg gctgcccttc tttgacgtat aatcttctaa ttaatacccg
     6841 gccttgtcaa agtggagcac aaacgttaat taattcccca gcaggcaggt aattaacagt
     6901 gtgactccct ttttgctgcg agtggggctg atacagagag atgtggcact atggagccca
     6961 cggggtcctg gcactgggtg cccacggagg tccccatgtg ctgcagtgtc accgcctccg
     7021 aggagacagt attgtccctg cggtgtccct gcagctcagc tctgtccaca gggccacctc
     7081 cagtttggag gggacacaat gcagccccga tgcaacccat cctcgcagca tcccagggac
     7141 aaagacccca ctgcaagacc gcacacaggg ctgggtcccg ctcccctaat atctacagtg
     7201 cttcttgcat ggccccttaa tcaatgcagt taatcagcat gcgctcatgc accgctctgg
     7261 agctgcaaag cccctcgcag cgctgctcac caacaccgcg caccgccccg gcccagcctg
     7321 cagcacgcgc tgcaaacagg aaagaaacaa aatattgccc aaatgtaggc aaaggcattc
     7381 ggctgccttg acctccgccg ggccgggccc tgcctgactc agctccttac tcagcgctcg
     7441 cttcctccct ccggctgcca ccgccgcagc gcacaccctg acaaagagtg gcccttaacg
     7501 ggctctgagg tgcacccagc agtgcactca gcagtccaag ggccggcctg gaggtttgca
     7561 ccgctacgtg ctgacattag cattgaactt ggccctgggt agtgctgcag gccgggcggg
     7621 gtgggtgtag agagtgcagc gcgcgttgca cccggtgccc cttcccctcc cttgcatccc
     7681 agcaggctgc accccagcac caggcccgtg catgcatgct cctggtgtta ttgcagcctg
     7741 gtgcatgcat gcgtcttgat ggtgcagcgc tgtgcatgca tcctccttgg tgtgtagcag
     7801 tctagtgcat gcatacccct cggtgttatt gctgctctgt gcacgcacgc tcattgtatc
     7861 acttcatccc agtgcatgca ctcacactgg agcgattgct gctcggtgca cgcacactca
     7921 ttgtatcact gcagctcagt ggctgcacgc acaccggtgt tattgctgct cggtgcgtgc
     7981 atgcacatca gtgtcgctgc agctcagtgc atgcacgctc attgcccatc gctatccctg
     8041 cctctcctgc tggcgctccc cgggaggtga cttcaagggg accgcaggac cacctcgggg
     8101 gtggggggag ggctgcacac gcggaccccg ctccccctcc ccaacaaagc actgtggaat
     8161 caaaaagggg ggagggggga tggaggggcg cgtcacaccc ccgccccaca ccctcacctc
     8221 gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc acccccaatt
     8281 ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg
//
