LOCUS       sequence-346988-        8381 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutively active KIF1A for optogenetic heterodimerization to 
            iLID via SSPB(nano).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8381)
  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 8381)
  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(1-383)-GFP-SSPB(nano)
FEATURES             Location/Qualifiers
     source          1..8381
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1057..1079
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     regulatory      1142..1151
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2124..2133
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2396..2398,2399..3109)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2396..2398,2399..3109)
                     /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(2438..2459)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2699..2718)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3046..3067
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3239..3568
                     /codon_start=1
                     /product="SspB from Haemophilus influenzae, with the 
                     mutations Y11K and A15E to disrupt the natural dimerization
                     of the protein"
                     /label=SspB nano
                     /note="dimerizes with iLID at high affinity to create iLID 
                     nano (Guntas et al., 2015)"
                     /translation="SSPKRPKLLREYYDWLVDNSFTPYLVVDATYLGVNVPVEYVKDGQ
                     IVLNLSASATGNLQLTNDFIQFNARFKGVSRELYIPMGAALAIYARENGDGVMFEPEEI
                     YDELNI"
     polyA_signal    3701..3835
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3738..3757)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3792..3811
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3849..3865)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3849..3865)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3862..3884)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3873..3889
                     /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(3897..3903,3904..3921,3922..3927))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3942..3963
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4080..4097)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4251..4839)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4331..4350)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5010..5801,5802..5870))
                     /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     5633..5652
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5871..5975)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6043..6061
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6099..6121)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6221..6240
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6434..6456
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6448..6465
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6449..6465
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gcgaggcgga gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa gtttcctttt
       61 atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg ggcgggagtc
      121 gctgcgcgct gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg cccgccccgg
      181 ctctgactga ccgcgttact cccacaggtg agcgggcggg acggcccttc tcctccgggc
      241 tgtaattagc gcttggttta atgacggctc gtttcttttc tgtggctgcg tgaaagcctt
      301 aaagggctcc gggagggccc tttgtgcggg ggggagcggc tcggggggtg cgtgcgtgtg
      361 tgtgtgcgtg gggagcgccg cgtgcggctc cgcgctgccc ggcggctgtg agcgctgcgg
      421 gcgcggcgcg gggctttgtg cgctccgcag tgtgcgcgag gggagcgcgg ccgggggcgg
      481 tgccccgcgg tgcggggggg gctgcgaggg gaacaaaggc tgcgtgcggg gtgtgtgcgt
      541 gggggggtga gcagggggtg tgggcgcggc ggtcgggctg taaccccccc ctgcaccccc
      601 ctccccgagt tgctgagcac ggcccggctt cgggtgcggg gctccgtgcg gggcgtggcg
      661 cggggctcgc cgtgccgggc ggggggtggc ggcaggtggg ggtgccgggc ggggcggggc
      721 cgcctcgggc cggggagggc tcgggggagg ggcgcggcgg cccccggagc gccggcggct
      781 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
      841 gacttccttt gtcccaaatc tgtgcggagc cgaaatctgg gaggcgccgc cgcaccccct
      901 ctagcgggcg cggggcgaag cggtgcggcg ccggcaggaa ggaaatgggc ggggagggcc
      961 ttcgtgcgtc gccgcgccgc cgtccccttc tccatctcca gcctcggggc tgtccgcagg
     1021 gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg
     1081 gcggggttta tatcttccct tctctgttcc tccgcagccc ccaagctagg caattgggcg
     1141 cgccaccatg gccggggctt cggtgaaggt ggcggtgcgt gtccgcccct tcaactcccg
     1201 ggaaatgagc cgagactcca agtgcatcat tcagatgtct ggaagcacca ccactattgt
     1261 caaccccaaa cagcccaagg agacacccaa gagcttcagc tttgattact cctactggtc
     1321 acatacctcg ccggaggaca tcaactatgc ctctcagaag caggtctaca gggacattgg
     1381 ggaagagatg ctacagcatg cctttgaggg ctacaatgtg tgcatctttg cctatgggca
     1441 gacgggggcc ggcaagtcct acaccatgat ggggaaacag gagaaggacc agcaaggcat
     1501 cattccacag ctctgcgaag acctcttctc tcgaatcaac gacaccacca acgacaacat
     1561 gtcctactct gtggaggtca gctacatgga gatttactgt gagcgagtcc gtgacctcct
     1621 gaaccccaag aacaaaggca acctgcgtgt gagggagcac cctctgctgg ggccttatgt
     1681 ggaggacctc tccaagctgg ctgtaacctc ctacaatgac atccaggacc tcatggactc
     1741 agggaacaag gccaggactg tggcggccac aaacatgaat gagaccagca gccgttctca
     1801 cgcagtcttc aacatcatct ttacccagaa gcgccatgat gcagagacca acatcaccac
     1861 agagaaagtg agcaaaatca gcctggtgga cctggcaggc agtgaacgag ccgactccac
     1921 aggagccaag ggcacgcgac tcaaggaggg ggccaacatc aacaagtctt tgaccaccct
     1981 gggaaaggtc atctctgctc tggctgaaat ggactctgga cccaacaaga acaagaaaaa
     2041 gaagaagaca gatttcatcc cttaccgaga ctccgtattg acatggcttc tccgggaaaa
     2101 cctcggtggc aactcaagga cagccatggt ggcagccctg agccctgcgg acatcaacta
     2161 tgatgagacc ctcagcacac tgagatatgc agatcgggcc aaacaaatcc gctgcaatgc
     2221 catcatcaat gaagacccca acaacaagct gattagagaa ctgaaggatg aggtgacccg
     2281 gctgcgggac cttctgtcta gaggtggcgg gggcagtggg ggagggggaa gtggaggcgg
     2341 cggttctggc gggggaggct ccggtggcgg ggggagcggg ggtgggagta agcttgtgag
     2401 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt
     2461 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct
     2521 gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac
     2581 caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga
     2641 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga
     2701 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg
     2761 cattgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga
     2821 gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa
     2881 ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta
     2941 ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag
     3001 cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga
     3061 gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagg ctagcggagg
     3121 cggtggctca ggcggaggag ggtcaggagg gggaggcagt ggcgggggcg gctctggagg
     3181 agggggttct ggtggtggta gctacgtaga tctgagcggc ctgaccctgc aggaattcag
     3241 ctccccgaaa cgccctaagc tgctgcgtga atattacgat tggctggttg ataacagctt
     3301 taccccatat ctggtggtgg atgccacata cctgggcgtg aacgtgcccg tggagtatgt
     3361 gaaagacggt cagatcgtgc tgaatctgtc tgcaagtgcg accggcaacc tgcaactgac
     3421 aaatgatttt atccagttca acgcccgctt taagggcgtg tctcgtgaac tgtatatccc
     3481 gatgggtgcc gctctggcca tttacgctcg cgagaacggc gatggtgtga tgttcgaacc
     3541 agaagaaatc tatgacgagc tgaatattgg ttaaaccggt cctagataag taatgatcat
     3601 aatcagccat atcacatctg tagaggtttt acttgcttta aaaaacctcc cacacctccc
     3661 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
     3721 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     3781 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatcaattc
     3841 gtaatcatgt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     3901 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     3961 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     4021 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4081 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4141 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4201 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4261 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4321 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4381 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4441 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4501 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4561 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4621 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     4681 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     4741 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     4801 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4861 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4921 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4981 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5041 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5101 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5161 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5221 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5281 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5341 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5401 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     5461 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5521 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5581 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     5641 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5701 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5761 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5821 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5881 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5941 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6001 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
     6061 ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg
     6121 gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg
     6181 tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta
     6241 ctgagagtgc accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc
     6301 atcaggcgcc attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc
     6361 tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta
     6421 acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtgccaa gttgggatct
     6481 ttgcattggc ccacggctct caggatgggg atgctcccct tcagcacccg gttccccttg
     6541 gaaactgatg gtcctggctc tgtggcatgg cagtggcact gtgaggagcc ccctaccagc
     6601 agcacacagt ggggttggca ctgccacgct ccggatgccg cgctctgatc caaccccata
     6661 atcaagggaa cccgaattgc cccatcattg cccccaccac ccccatcctg ccgggccctc
     6721 acaccccacg ctgccttctg gtgacattcc ccagcccaaa cccacggctt catggctacc
     6781 gcggggcatt tcccattgcc gccccattat cagctctgca cacctcccgc tgtacccatg
     6841 cctcgtggct gcccttcttt gacgtataat cttctaatta atacccggcc ttgtcaaagt
     6901 ggagcacaaa cgttaattaa ttccccagca ggcaggtaat taacagtgtg actccctttt
     6961 tgctgcgagt ggggctgata cagagagatg tggcactatg gagcccacgg ggtcctggca
     7021 ctgggtgccc acggaggtcc ccatgtgctg cagtgtcacc gcctccgagg agacagtatt
     7081 gtccctgcgg tgtccctgca gctcagctct gtccacaggg ccacctccag tttggagggg
     7141 acacaatgca gccccgatgc aacccatcct cgcagcatcc cagggacaaa gaccccactg
     7201 caagaccgca cacagggctg ggtcccgctc ccctaatatc tacagtgctt cttgcatggc
     7261 cccttaatca atgcagttaa tcagcatgcg ctcatgcacc gatctggagc tgcaaagccc
     7321 ctcgcagcgc tgctcaccaa caccgcgcac cgccccggcc cagcctgcag cacgcgctgc
     7381 aaacaggaaa gaaacaaaat attgcccaaa tgtaggcaaa ggcattcggc tgccttgacc
     7441 tccgccgggc cgggccctgc ctgactcagc tccttactca gcgctcgctt cctccctccg
     7501 gctgccaccg ccgcagcgca caccctgaca aagagtggcc cttaacgggc tctgaggtgc
     7561 acccagcagt gcactcagca gtccaagggc cggcctggag gtttgcaccg ctacgtgctg
     7621 acattagcat tgaacttggc cctgggtagt gctgcaggcc gggcggggtg ggtgtagaga
     7681 gtgcagcgcg cgttgcaccc ggtgcccctt cccctccctt gcatcccagc aggctgcacc
     7741 ccagcaccag gcccgtgcat gcatgctcct ggtgttattg cagcctggtg catgcatgcg
     7801 tcttgatggt gcagcgctgt gcatgcatcc tccttggtgt gtagcagtct agtgcatgca
     7861 tacccctcgg tgttattgct gctctgtgca cgcacgctca ttgtatcact tcatcccagt
     7921 gcatgcactc acactggagc gattgctgct cggtgcacgc acactcattg tatcactgca
     7981 gctcagtggc tgcacgcaca ccggtgttat tgctgctcgg tgcgtgcatg cacatcagtg
     8041 tcgctgcagc tcagtgcatg cacgctcatt gcccatcgct atccctgcct ctcctgctgg
     8101 cgctccccgg gaggtgactt caaggggacc gcaggaccac ctcgggggtg gggggagggc
     8161 tgcacacgcg gaccccgctc cccctcccca acaaagcact gtggaatcaa aaagggggga
     8221 ggggggatgg aggggcgcgt cacacccccg ccccacaccc tcacctcgag gtgagcccca
     8281 cgttctgctt cactctcccc atctcccccc cctccccacc cccaattttg tatttattta
     8341 ttttttaatt attttgtgca gcgatggggg cggggggggg g
//
