LOCUS       Exported                8769 bp ds-DNA     circular SYN 04-NOV-2021
DEFINITION  Optogenetic coupling to tetramerized moss kinesin-14 via SSPB(micro)
            for highly efficient retrograde transport .
ACCESSION   .
VERSION     .
KEYWORDS    pB80-SSPB(micro)-GFP-GCN4-ppKin14VIb(861-1321)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8769)
  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 8769)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 4, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8769
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     941..963
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     regulatory      1026..1035
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1503..2216
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             1503..2216
                     /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(1545..1566)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1806..1825)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2153..2174
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    2226..2250
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     RBS             2714..2722
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     polyA_signal    3927..4061
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3964..3983)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4018..4037
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4075..4091)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4075..4091)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4088..4110)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4099..4115
                     /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(4123..4153)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4168..4189
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4306..4323)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4477..5065)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4557..4576)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5236..6096)
                     /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     5859..5878
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6097..6201)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6269..6287
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6325..6347)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6447..6466
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6660..6682
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6674..6691
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6675..6691
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        8492..8767
                     /label=chicken beta-actin promoter
ORIGIN
        1 agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc
       61 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      121 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      181 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      241 tgtgtgtgtg cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc tgtgagcgct
      301 gcgggcgcgg cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc gcggccgggg
      361 gcggtgcccc gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg cggggtgtgt
      421 gcgtgggggg gtgagcaggg ggtgtgggcg cggcggtcgg gctgtaaccc ccccctgcac
      481 ccccctcccc gagttgctga gcacggcccg gcttcgggtg cggggctccg tgcggggcgt
      541 ggcgcggggc tcgccgtgcc gggcgggggg tggcggcagg tgggggtgcc gggcggggcg
      601 gggccgcctc gggccgggga gggctcgggg gaggggcgcg gcggcccccg gagcgccggc
      661 ggctgtcgag gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg cgagagggcg
      721 cagggacttc ctttgtccca aatctgtgcg gagccgaaat ctgggaggcg ccgccgcacc
      781 ccctctagcg ggcgcggggc gaagcggtgc ggcgccggca ggaaggaaat gggcggggag
      841 ggccttcgtg cgtcgccgcg ccgccgtccc cttctccatc tccagcctcg gggctgtccg
      901 cagggggacg gctgccttcg ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg
      961 accggcgggg tttatatctt cccttctctg ttcctccgca gcccccaagc taggcaattg
     1021 ggcgcgccac catggatctg agcggcctga ccctgcagga attcagctcc ccgaaacgcc
     1081 ctaagctgct gcgtgaatat tacgattggc tggttgataa cagctttacc ccatatctgg
     1141 tggtggatgc cacatacctg ggcgtgaacg tgcccgtgga gtatgtgaaa gacggtcaga
     1201 tcgtgctgaa tctgtctgca agtgcgaccg gcaacctgca actgacaaat gattttatcc
     1261 agttcaacgc ccagtttaag ggcgtgtctc gtgaactgta tatcccgatg ggtgccgctc
     1321 tggccattta cgctcgcgag aacggcgatg gtgtgatgtt cgaaccagaa gaaatctatg
     1381 acgagctgaa tattggtcaa ttgtctagag gtggcggggg cagtggggga gggggaagtg
     1441 gaggcggcgg ttctggcggg ggaggctccg gkggcggggg grgsgggggk gggagtaagc
     1501 ttgtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     1561 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     1621 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     1681 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     1741 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     1801 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     1861 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     1921 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     1981 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     2041 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     2101 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     2161 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagcacc
     2221 ggtatacaag tttgtacaaa aaagcaggct ccgctattga agacaagttg gaagaaatac
     2281 tttcgaaatt gtaccacatt gaaaatgagt tggccagaat taagaaatta ttgggcgaaa
     2341 ttggtggagg tagtggtgga ggtagtgcgg ccgccagaga tgatttgagg ggacgacttg
     2401 taactctcga gaagaaatta gcagagcgta cccaagattt gagtcttgca gagtctactt
     2461 tggcaactcg tgagggtgaa ttcgaatccc tgcaaagcaa tctaatggag cttaatgagc
     2521 tccgggagat gaaggaggat atcgacagaa agaatgagca aacagcagct atattgaagc
     2581 gacaagcaga ccagataact gaattagaaa ctctttatag ggaagaacaa gtccttcgaa
     2641 aacgatactt caacatgatg gaagacatga aaggtaagat cagggtatat gctcggtgga
     2701 gacccctaag tagtaaggag gtcaaagagc ggcaacaaaa cgtgttgata gcacctgatg
     2761 aattcaccat cgaacatcct tggaaagacg acaaacccaa acaacatcaa tttgatcacg
     2821 ttttcgacca ccatgcgacc caagaagagg tctttgagga tacaaagtat ttggtccaat
     2881 cagcaattga cggatacaat gtctgtatat ttgcatacgg gcaaactggg tcaggcaaaa
     2941 cgtttacgat ttacggttct gacaacaacc caggtctaac tccacgagct accaaagaac
     3001 tttttggcta tcttaagcga gacgccaata agttttcgtt tgctctgaag gtttacatgc
     3061 ttgaaatata tcaagattcg ctcatcgatc tgcttctacc aaagagtgct gcaaagccac
     3121 gaaagcttga gattaagaag gactccaagg gaatggttgt cgtagagaat gcaactctac
     3181 tgccgattgc atcccatgat gaactgcaag ccattgttca taagggtctg gagagacgac
     3241 atgtttcggg cacccacatg aatgcggaga gttcacggtc acacctcatt ctatcagtaa
     3301 tcgttgaaag caccaaccga cagtctcaag ttcttgtgaa gggcaagctg agttttgtgg
     3361 atcttgcggg ctctgaaaga gtgaagaaat ctggatccag tggtgaacaa ctgaaagagg
     3421 cccaaagcat taataaatct ttgtcagcac taggtgatgt cattagcgcc cttgcaacag
     3481 aggagcagca tattccttac cgtaaccaca agctaacaat gcttatgagc gattctctcg
     3541 gtggaaatgc gaagaccctc atgtttgtaa atatctcacc agcagagtct aatctggacg
     3601 agacccataa ttcactctgc tatgccacaa gggtaagatc catcatcaat gaggccagta
     3661 aaaatactac aacgaaagag atactgcgac tgaagaaaca aatcgcgtac tggaaggagc
     3721 aagcgggcag gatgggaggg tccgaggatc tggaagaggt tatggatgat cgtggaatca
     3781 aaactgcaga ttaaacgcgt accggtccta gataagtaat gatcataatc agccatatca
     3841 catctgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac
     3901 ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat
     3961 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     4021 gtttgtccaa actcatcaat gtatcttatc atgtctggat caattcgtaa tcatgtcata
     4081 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     4141 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     4201 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     4261 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     4321 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     4381 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     4441 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     4501 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     4561 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     4621 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     4681 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     4741 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     4801 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     4861 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     4921 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     4981 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     5041 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     5101 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     5161 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     5221 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     5281 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     5341 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     5401 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     5461 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     5521 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     5581 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     5641 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     5701 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     5761 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     5821 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     5881 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     5941 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     6001 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     6061 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     6121 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     6181 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     6241 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc
     6301 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     6361 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     6421 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     6481 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc
     6541 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     6601 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     6661 ccagtcacga cgttgtaaaa cgacggccag tgccaagttg ggatctttgc attggcccac
     6721 ggctctcagg atggggatgc tccccttcag cacccggttc cccttggaaa ctgatggtcc
     6781 tggctctgtg gcatggcagt ggcactgtga ggagccccct accagcagca cacagtgggg
     6841 ttggcactgc cacgctccgg atgccgcgct ctgatccaac cccataatca agggaacccg
     6901 aattgcccca tcattgcccc caccaccccc atcctgccgg gccctcacac cccacgctgc
     6961 cttctggtga cattccccag cccaaaccca cggcttcatg gctaccgcgg ggcatttccc
     7021 attgccgccc cattatcagc tctgcacacc tcccgctgta cccatgcctc gtggctgccc
     7081 ttctttgacg tataatcttc taattaatac ccggccttgt caaagtggag cacaaacgtt
     7141 aattaattcc ccagcaggca ggtaattaac agtgtgactc cctttttgct gcgagtgggg
     7201 ctgatacaga gagatgtggc actatggagc ccacggggtc ctggcactgg gtgcccacgg
     7261 aggtccccat gtgctgcagt gtcaccgcct ccgaggagac agtattgtcc ctgcggtgtc
     7321 cctgcagctc agctctgtcc acagggccac ctccagtttg gaggggacac aatgcagccc
     7381 cgatgcaacc catcctcgca gcatcccagg gacaaagacc ccactgcaag accgcacaca
     7441 gggctgggtc ccgctcccct aatatctaca gtgcttcttg catggcccct taatcaatgc
     7501 agttaatcag catgcgctca tgcaccgctc tggagctgca aagcccctcg cagcgctgct
     7561 caccaacacc gcgcaccgcc ccggcccagc ctgcagcacg cgctgcaaac aggaaagaaa
     7621 caaaatattg cccaaatgta ggcaaaggca ttcggctgcc ttgacctccg ccgggccggg
     7681 ccctgcctga ctcagctcct tactcagcgc tcgcttcctc cctccggctg ccaccgccgc
     7741 agcgcacacc ctgacaaaga gtggccctta acgggctctg aggtgcaccc agcagtgcac
     7801 tcagcagtcc aagggccggc ctggaggttt gcaccgctac gtgctgacat tagcattgaa
     7861 cttggccctg ggtagtgctg caggccgggc ggggtgggtg tagagagtgc agcgcgcgtt
     7921 gcacccggtg ccccttcccc tcccttgcat cccagcaggc tgcaccccag caccaggccc
     7981 gtgcatgcat gctcctggtg ttattgcagc ctggtgcatg catgcgtctt gatggtgcag
     8041 cgctgtgcat gcatcctcct tggtgtgtag cagtctagtg catgcatacc cctcggtgtt
     8101 attgctgctc tgtgcacgca cgctcattgt atcacttcat cccagtgcat gcactcacac
     8161 tggagcgatt gctgctcggt gcacgcacac tcattgtatc actgcagctc agtggctgca
     8221 cgcacaccgg tgttattgct gctcggtgcg tgcatgcaca tcagtgtcgc tgcagctcag
     8281 tgcatgcacg ctcattgccc atcgctatcc ctgcctctcc tgctggcgct ccccgggagg
     8341 tgacttcaag gggaccgcag gaccacctcg ggggtggggg gagggctgca cacgcggacc
     8401 ccgctccccc tccccaacaa agcactgtgg aatcaaaaag gggggagggg ggatggaggg
     8461 gcgcgtcaca cccccgcccc acaccctcac ctcgaggtga gccccacgtt ctgcttcact
     8521 ctccccatct cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt
     8581 tgtgcagcga tgggggcggg gggggggggg gcgcgcgcca ggcggggcgg ggcggggcgr
     8641 ggggcggggc ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg
     8701 aaagtttcct tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg
     8761 gcgggcggg
//
