LOCUS       sequence-346982-        8735 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Opto-kinesin for optogenetic heterodimerization to iLID via 
            SSPB(micro).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8735)
  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 8735)
  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)-VVDfast-GFP-SSPB(micro)
FEATURES             Location/Qualifiers
     source          1..8735
                     /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(2754..2756,2757..3467)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2754..2756,2757..3467)
                     /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(2796..2817)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3057..3076)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3404..3425
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             join(3597..3803,3804..3806,3807..3926)
                     /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    4059..4193
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4096..4115)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4150..4169
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4207..4223)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4207..4223)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4220..4242)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4231..4247
                     /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(4255..4261,4262..4279,4280..4285))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4300..4321
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4438..4455)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4609..5197)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4689..4708)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5368..6159,6160..6228))
                     /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     5991..6010
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6229..6333)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6401..6419
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6457..6479)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6579..6598
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6792..6814
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6806..6823
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6807..6823
                     /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 gggggatccc
     2281 atactcttta tgcccccggt ggatatgaca ttatgggata tctgtgccag atcatgaacc
     2341 ggccaaaccc gcaggtggaa ctgggccccg tggatacatc ctgcgccttg gttctttgtg
     2401 acctgaaaca gaaagacacc ccggtagttt acgcgagtga agccttcctc tacatgacag
     2461 gttacagcaa cgcagaggtg ctgggccgga attgccggtt tctgcaaagc cctgacggca
     2521 tggtgaagcc caagagcacc cggaagtacg tggatagtaa cacaatcaat actatgcgca
     2581 aggcaatcga caggaatgcc gaggtgcagg ttgaagtagt caattttaaa aagaatggac
     2641 agcgatttgt taatttcctg actatgatac ctgttaggga cgaaacaggc gagtatcgat
     2701 actctatggg attccagtgc gaagtcgacg ggagcggggg tgggagtaag cttgtgagca
     2761 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
     2821 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
     2881 ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
     2941 ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact
     3001 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
     3061 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
     3121 ttgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
     3181 acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg
     3241 tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc
     3301 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca
     3361 cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
     3421 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaaggct agcggaggcg
     3481 gtggctcagg cggaggaggg tcaggagggg gaggcagtgg cgggggcggc tctggaggag
     3541 ggggttctgg tggtggtagc tacgtagatc tgagcggcct gaccctgcag gaattcagct
     3601 ccccgaaacg ccctaagctg ctgcgtgaat attacgattg gctggttgat aacagcttta
     3661 ccccatatct ggtggtggat gccacatacc tgggcgtgaa cgtgcccgtg gagtatgtga
     3721 aagacggtca gatcgtgctg aatctgtctg caagtgcgac cggcaacctg caactgacaa
     3781 atgattttat ccagttcaac gcccagttta agggcgtgtc tcgtgaactg tatatcccga
     3841 tgggtgccgc tctggccatt tacgctcgcg agaacggcga tggtgtgatg ttcgaaccag
     3901 aagaaatcta tgacgagctg aatattggtt aaaccggtcc tagataagta atgatcataa
     3961 tcagccatat cacatctgta gaggttttac ttgctttaaa aaacctccca cacctccccc
     4021 tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata
     4081 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     4141 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg atcaattcgt
     4201 aatcatgtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     4261 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     4321 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     4381 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     4441 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     4501 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     4561 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     4621 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     4681 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     4741 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     4801 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     4861 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     4921 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     4981 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     5041 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     5101 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     5161 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     5221 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     5281 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     5341 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     5401 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     5461 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     5521 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     5581 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     5641 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     5701 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     5761 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     5821 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     5881 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     5941 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     6001 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     6061 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     6121 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     6181 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     6241 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     6301 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     6361 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
     6421 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     6481 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     6541 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
     6601 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     6661 caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     6721 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
     6781 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgccaagt tgggatcttt
     6841 gcattggccc acggctctca ggatggggat gctccccttc agcacccggt tccccttgga
     6901 aactgatggt cctggctctg tggcatggca gtggcactgt gaggagcccc ctaccagcag
     6961 cacacagtgg ggttggcact gccacgctcc ggatgccgcg ctctgatcca accccataat
     7021 caagggaacc cgaattgccc catcattgcc cccaccaccc ccatcctgcc gggccctcac
     7081 accccacgct gccttctggt gacattcccc agcccaaacc cacggcttca tggctaccgc
     7141 ggggcatttc ccattgccgc cccattatca gctctgcaca cctcccgctg tacccatgcc
     7201 tcgtggctgc ccttctttga cgtataatct tctaattaat acccggcctt gtcaaagtgg
     7261 agcacaaacg ttaattaatt ccccagcagg caggtaatta acagtgtgac tccctttttg
     7321 ctgcgagtgg ggctgataca gagagatgtg gcactatgga gcccacgggg tcctggcact
     7381 gggtgcccac ggaggtcccc atgtgctgca gtgtcaccgc ctccgaggag acagtattgt
     7441 ccctgcggtg tccctgcagc tcagctctgt ccacagggcc acctccagtt tggaggggac
     7501 acaatgcagc cccgatgcaa cccatcctcg cagcatccca gggacaaaga ccccactgca
     7561 agaccgcaca cagggctggg tcccgctccc ctaatatcta cagtgcttct tgcatggccc
     7621 cttaatcaat gcagttaatc agcatgcgct catgcaccgc tctggagctg caaagcccct
     7681 cgcagcgctg ctcaccaaca ccgcgcaccg ccccggccca gcctgcagca cgcgctgcaa
     7741 acaggaaaga aacaaaatat tgcccaaatg taggcaaagg cattcggctg ccttgacctc
     7801 cgccgggccg ggccctgcct gactcagctc cttactcagc gctcgcttcc tccctccggc
     7861 tgccaccgcc gcagcgcaca ccctgacaaa gagtggccct taacgggctc tgaggtgcac
     7921 ccagcagtgc actcagcagt ccaagggccg gcctggaggt ttgcaccgct acgtgctgac
     7981 attagcattg aacttggccc tgggtagtgc tgcaggccgg gcggggtggg tgtagagagt
     8041 gcagcgcgcg ttgcacccgg tgccccttcc cctcccttgc atcccagcag gctgcacccc
     8101 agcaccaggc ccgtgcatgc atgctcctgg tgttattgca gcctggtgca tgcatgcgtc
     8161 ttgatggtgc agcgctgtgc atgcatcctc cttggtgtgt agcagtctag tgcatgcata
     8221 cccctcggtg ttattgctgc tctgtgcacg cacgctcatt gtatcacttc atcccagtgc
     8281 atgcactcac actggagcga ttgctgctcg gtgcacgcac actcattgta tcactgcagc
     8341 tcagtggctg cacgcacacc ggtgttattg ctgctcggtg cgtgcatgca catcagtgtc
     8401 gctgcagctc agtgcatgca cgctcattgc ccatcgctat ccctgcctct cctgctggcg
     8461 ctccccggga ggtgacttca aggggaccgc aggaccacct cgggggtggg gggagggctg
     8521 cacacgcgga ccccgctccc cctccccaac aaagcactgt ggaatcaaaa aggggggagg
     8581 ggggatggag gggcgcgtca cacccccgcc ccacaccctc acctcgaggt gagccccacg
     8641 ttctgcttca ctctccccat ctcccccccc tccccacccc caattttgta tttatttatt
     8701 ttttaattat tttgtgcagc gatgggggcg ggggg
//
