LOCUS       sequence-346991-        8713 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 8713)
  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 8713)
  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-mCherry-SSPB(micro)
FEATURES             Location/Qualifiers
     source          1..8713
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1047..1069
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     regulatory      1132..1141
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2114..2123
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2737..3441
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(2879..2897)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             join(3571..3777,3778..3780,3781..3900)
                     /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    4033..4167
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4070..4089)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4124..4143
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4181..4197)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4181..4197)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4194..4216)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4205..4221
                     /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(4229..4235,4236..4253,4254..4259))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4274..4295
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4412..4429)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4583..5171)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4663..4682)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5342..6133,6134..6202))
                     /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     5965..5984
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6203..6307)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6375..6393
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6431..6453)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6553..6572
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6766..6788
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6780..6797
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6781..6797
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa gtttcctttt atggcgaggc
       61 ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg ggcgggagtc gctgcgcgct
      121 gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg cccgccccgg ctctgactga
      181 ccgcgttact cccacaggtg agcgggcggg acggcccttc tcctccgggc tgtaattagc
      241 gcttggttta atgacggctc gtttcttttc tgtggctgcg tgaaagcctt aaagggctcc
      301 gggagggccc tttgtgcggg ggggagcggc tcggggggtg cgtgcgtgtg tgtgtgcgtg
      361 gggagcgccg cgtgcggctc cgcgctgccc ggcggctgtg agcgctgcgg gcgcggcgcg
      421 gggctttgtg cgctccgcag tgtgcgcgag gggagcgcgg ccgggggcgg tgccccgcgg
      481 tgcggggggg gctgcgaggg gaacaaaggc tgcgtgcggg gtgtgtgcgt gggggggtga
      541 gcagggggtg tgggcgcggc ggtcgggctg taaccccccc ctgcaccccc ctccccgagt
      601 tgctgagcac ggcccggctt cgggtgcggg gctccgtgcg gggcgtggcg cggggctcgc
      661 cgtgccgggc ggggggtggc ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc
      721 cggggagggc tcgggggagg ggcgcggcgg cccccggagc gccggcggct gtcgaggcgc
      781 ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg gacttccttt
      841 gtcccaaatc tgtgcggagc cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg
      901 cggggcgaag cggtgcggcg ccggcaggaa ggaaatgggc ggggagggcc ttcgtgcgtc
      961 gccgcgccgc cgtccccttc tccatctcca gcctcggggc tgtccgcagg gggacggctg
     1021 ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg gcggggttta
     1081 tatcttccct tctctgttcc tccgcagccc ccaagctagg caattgggcg cgccaccatg
     1141 gccggggctt cggtgaaggt ggcggtgcgt gtccgcccct tcaactcccg ggaaatgagc
     1201 cgagactcca agtgcatcat tcagatgtct ggaagcacca ccactattgt caaccccaaa
     1261 cagcccaagg agacacccaa gagcttcagc tttgattact cctactggtc acatacctcg
     1321 ccggaggaca tcaactatgc ctctcagaag caggtctaca gggacattgg ggaagagatg
     1381 ctacagcatg cctttgaggg ctacaatgtg tgcatctttg cctatgggca gacgggggcc
     1441 ggcaagtcct acaccatgat ggggaaacag gagaaggacc agcaaggcat cattccacag
     1501 ctctgcgaag acctcttctc tcgaatcaac gacaccacca acgacaacat gtcctactct
     1561 gtggaggtca gctacatgga gatttactgt gagcgagtcc gtgacctcct gaaccccaag
     1621 aacaaaggca acctgcgtgt gagggagcac cctctgctgg ggccttatgt ggaggacctc
     1681 tccaagctgg ctgtaacctc ctacaatgac atccaggacc tcatggactc agggaacaag
     1741 gccaggactg tggcggccac aaacatgaat gagaccagca gccgttctca cgcagtcttc
     1801 aacatcatct ttacccagaa gcgccatgat gcagagacca acatcaccac agagaaagtg
     1861 agcaaaatca gcctggtgga cctggcaggc agtgaacgag ccgactccac aggagccaag
     1921 ggcacgcgac tcaaggaggg ggccaacatc aacaagtctt tgaccaccct gggaaaggtc
     1981 atctctgctc tggctgaaat ggactctgga cccaacaaga acaagaaaaa gaagaagaca
     2041 gatttcatcc cttaccgaga ctccgtattg acatggcttc tccgggaaaa cctcggtggc
     2101 aactcaagga cagccatggt ggcagccctg agccctgcgg acatcaacta tgatgagacc
     2161 ctcagcacac tgagatatgc agatcgggcc aaacaaatcc gctgcaatgc catcatcaat
     2221 gaagacccca actctagagg tggcgggggc agtgggggat cccatactct ttatgccccc
     2281 ggtggatatg acattatggg atatctgtgc cagatcatga accggccaaa cccgcaggtg
     2341 gaactgggcc ccgtggatac atcctgcgcc ttggttcttt gtgacctgaa acagaaagac
     2401 accccggtag tttacgcgag tgaagccttc ctctacatga caggttacag caacgcagag
     2461 gtgctgggcc ggaattgccg gtttctgcaa agccctgacg gcatggtgaa gcccaagagc
     2521 acccggaagt acgtggatag taacacaatc aatactatgc gcaaggcaat cgacaggaat
     2581 gccgaggtgc aggttgaagt agtcaatttt aaaaagaatg gacagcgatt tgttaatttc
     2641 ctgactatga tacctgttag ggacgaaaca ggcgagtatc gatactctat gggattccag
     2701 tgcgaagtcg acgggagcgg gggtgggagt aagcttgtga gcaagggcga ggaggataac
     2761 atggccatca tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc
     2821 cacgagttcg agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc
     2881 aagctgaagg tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag
     2941 ttcatgtacg gctccaaggc ctacgtgaag caccccgccg acatccccga ctacttgaag
     3001 ctgtccttcc ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgtg
     3061 gtgaccgtga cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg
     3121 cgcggcacca acttcccctc cgacggtccc gtaatgcaga agaagaccat gggctgggag
     3181 gcctcctccg agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg
     3241 ctgaagctga aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag
     3301 aagcccgtgc agctgcccgg cgcctacaac gtcaacatca agttggacat cacctcccac
     3361 aacgaggact acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc
     3421 ggcatggacg agctgtacaa ggctagcgga ggcggtggct caggcggagg agggtcagga
     3481 gggggaggca gtggcggggg cggctctgga ggagggggtt ctggtggtgg tagctacgta
     3541 gatctgagcg gcctgaccct gcaggaattc agctccccga aacgccctaa gctgctgcgt
     3601 gaatattacg attggctggt tgataacagc tttaccccat atctggtggt ggatgccaca
     3661 tacctgggcg tgaacgtgcc cgtggagtat gtgaaagacg gtcagatcgt gctgaatctg
     3721 tctgcaagtg cgaccggcaa cctgcaactg acaaatgatt ttatccagtt caacgcccag
     3781 tttaagggcg tgtctcgtga actgtatatc ccgatgggtg ccgctctggc catttacgct
     3841 cgcgagaacg gcgatggtgt gatgttcgaa ccagaagaaa tctatgacga gctgaatatt
     3901 ggttaaaccg gtcctagata agtaatgatc ataatcagcc atatcacatc tgtagaggtt
     3961 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     4021 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     4081 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     4141 atcaatgtat cttatcatgt ctggatcaat tcgtaatcat gtcatagctg tttcctgtgt
     4201 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     4261 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     4321 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     4381 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     4441 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     4501 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     4561 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     4621 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     4681 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     4741 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     4801 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     4861 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     4921 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     4981 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     5041 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     5101 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     5161 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     5221 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     5281 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     5341 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     5401 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     5461 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     5521 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     5581 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     5641 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     5701 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     5761 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     5821 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     5881 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     5941 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     6001 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     6061 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     6121 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     6181 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     6241 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     6301 ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga
     6361 cattaaccta taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg
     6421 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     6481 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct
     6541 ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa
     6601 taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc
     6661 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
     6721 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
     6781 gtaaaacgac ggccagtgcc aagttgggat ctttgcattg gcccacggct ctcaggatgg
     6841 ggatgctccc cttcagcacc cggttcccct tggaaactga tggtcctggc tctgtggcat
     6901 ggcagtggca ctgtgaggag ccccctacca gcagcacaca gtggggttgg cactgccacg
     6961 ctccggatgc cgcgctctga tccaacccca taatcaaggg aacccgaatt gccccatcat
     7021 tgcccccacc acccccatcc tgccgggccc tcacacccca cgctgccttc tggtgacatt
     7081 ccccagccca aacccacggc ttcatggcta ccgcggggca tttcccattg ccgccccatt
     7141 atcagctctg cacacctccc gctgtaccca tgcctcgtgg ctgcccttct ttgacgtata
     7201 atcttctaat taatacccgg ccttgtcaaa gtggagcaca aacgttaatt aattccccag
     7261 caggcaggta attaacagtg tgactccctt tttgctgcga gtggggctga tacagagaga
     7321 tgtggcacta tggagcccac ggggtcctgg cactgggtgc ccacggaggt ccccatgtgc
     7381 tgcagtgtca ccgcctccga ggagacagta ttgtccctgc ggtgtccctg cagctcagct
     7441 ctgtccacag ggccacctcc agtttggagg ggacacaatg cagccccgat gcaacccatc
     7501 ctcgcagcat cccagggaca aagaccccac tgcaagaccg cacacagggc tgggtcccgc
     7561 tcccctaata tctacagtgc ttcttgcatg gccccttaat caatgcagtt aatcagcatg
     7621 cgctcatgca ccgctctgga gctgcaaagc ccctcgcagc gctgctcacc aacaccgcgc
     7681 accgccccgg cccagcctgc agcacgcgct gcaaacagga aagaaacaaa atattgccca
     7741 aatgtaggca aaggcattcg gctgccttga cctccgccgg gccgggccct gcctgactca
     7801 gctccttact cagcgctcgc ttcctccctc cggctgccac cgccgcagcg cacaccctga
     7861 caaagagtgg cccttaacgg gctctgaggt gcacccagca gtgcactcag cagtccaagg
     7921 gccggcctgg aggtttgcac cgctacgtgc tgacattagc attgaacttg gccctgggta
     7981 gtgctgcagg ccgggcgggg tgggtgtaga gagtgcagcg cgcgttgcac ccggtgcccc
     8041 ttcccctccc ttgcatccca gcaggctgca ccccagcacc aggcccgtgc atgcatgctc
     8101 ctggtgttat tgcagcctgg tgcatgcatg cgtcttgatg gtgcagcgct gtgcatgcat
     8161 cctccttggt gtgtagcagt ctagtgcatg catacccctc ggtgttattg ctgctctgtg
     8221 cacgcacgct cattgtatca cttcatccca gtgcatgcac tcacactgga gcgattgctg
     8281 ctcggtgcac gcacactcat tgtatcactg cagctcagtg gctgcacgca caccggtgtt
     8341 attgctgctc ggtgcgtgca tgcacatcag tgtcgctgca gctcagtgca tgcacgctca
     8401 ttgcccatcg ctatccctgc ctctcctgct ggcgctcccc gggaggtgac ttcaagggga
     8461 ccgcaggacc acctcggggg tggggggagg gctgcacacg cggaccccgc tccccctccc
     8521 caacaaagca ctgtggaatc aaaaaggggg gaggggggat ggaggggcgc gtcacacccc
     8581 cgccccacac cctcacctcg aggtgagccc cacgttctgc ttcactctcc ccatctcccc
     8641 cccctcccca cccccaattt tgtatttatt tattttttaa ttattttgtg cagcgatggg
     8701 ggcggggggg ggg
//
