LOCUS       sequence-254580-        6836 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Phosphomimic mutant of Stathmin/OP18 FRET-sensor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6836)
  AUTHORS   Van Geel O, Cheung S, Gadella TWJ
  TITLE     Combining optogenetics with sensitive FRET imaging to monitor local 
            microtubule manipulations.
  JOURNAL   Sci Rep. 2020 Apr 7;10(1):6034. doi: 10.1038/s41598-020-62874-3.
  PUBMED    32265472
REFERENCE   2  (bases 1 to 6836)
  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: "Sci Rep."; 
            date: "2020-04-7"; volume: "10(1):6034. doi"; pages: " 
            10.1038/s41598-020-62874-3"
COMMENT     Sequence Label: ROPY-4E
FEATURES             Location/Qualifiers
     source          1..6836
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     16..38
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     100..119
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     promoter        161..279
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring 
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
     CDS             354..371
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             381..398
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     regulatory      417..426
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(423..425,426..428,429..1139)
                     /codon_start=1
                     /product="monomeric optimized variant of EYFP, identical to
                     mVenus but with L68V (Kremers et al., 2006)"
                     /label=SYFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(468..489)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(729..748)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1076..1097
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1617..2309
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="VSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGGA
                     VTVTQDTSLEDGTLIYKVKLIGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIKM
                     ALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRHS
                     TGGMDEPYK"
     primer_bind     complement(1747..1765)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2015..2034
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2406..2425)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2471..2526
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2525..2544)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      2614..2661
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     2674..3379
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           2762..3134
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(2791..3379)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(3388..3421)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(3589..4176)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3668..3687)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4189..5004)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4912..4931
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     misc_recomb     5247..6074
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     CDS             5247..5371
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5409..6014)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     enhancer        6142..6445
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6447..6649
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6600..6620
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 gacggggcag ggcggggttc ggcttctggc gtgtgaccgg cggctctaga gcctctgcta
       61 accatgttca tgccttcttc tttttcctac agctcctggg caacgtgctg gttattgtgc
      121 tgtctcatca ttttggcaaa gaattggatc ggaccgaaat caccacaatt cagcaaattg
      181 tgaacatcat cacgttcatc tttccctggt tgccaaaggc ccattttcct gtcagtaacg
      241 agaaggtcgc gaattcaggc gctttttaga ctggtcgtag ggagaccaca acggtttccc
      301 tctagaaata attttgttta actataagaa ggagatatac catgcggggt tctcatcatc
      361 accaccatca cagcagcggc ctggtgccgc gcggcagctc gctagcgcta ccggtcgcca
      421 ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
      481 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
      541 acggcaagct gaccctgaag ctgatctgca ccaccggcaa gctgcccgtg ccctggccca
      601 ccctcgtgac caccctgggc tacggcgtgc agtgcttcgc ccgctacccc gaccacatga
      661 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
      721 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
      781 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
      841 acaagctgga gtacaactac aacagccaca acgtctatat caccgccgac aagcagaaga
      901 acggcatcaa ggccaacttc aagatccgcc acaacatcga ggacggcggc gtgcagctcg
      961 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     1021 actacctgag ctaccagtcc aagctgagca aagaccccaa cgagaagcgc gatcacatgg
     1081 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt
     1141 ccggactcag atctacgatg gcatcttctg atattcaggt gaaagagctg gagaagcgcg
     1201 ctgaaggcca ggcttttgag ctgattctcg aacctcggtc aaaagaatct gtccccgatt
     1261 tcccccttga gcccccaaag aagaaggacc tttccctgga ggaaatccag aagaaattag
     1321 aagctgcaga agaaagacgc aaggaacatg aggcggaagt cttgaagcag ctcgcggaga
     1381 agcgggagca tgagaaggag gtgctccaga aagccatcga ggagaacaac aacttcagca
     1441 agatggcgga ggagaagctg acccacaaaa tggaggctaa caaagagaac cgggaggcgc
     1501 agatggcggc caagctggag cgcttgcgag agaaggacaa gcacgtggaa gaggtgcgga
     1561 agaacaaaga atccaaagac cccgcggatg agactgaggc tgacggatcc ggttcagtga
     1621 gcaagggcga ggcagtgatc aaggagttca tgcggttcaa ggtgcacatg gagggctcca
     1681 tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc
     1741 agaccgccaa gctgaaggtg accaagggtg gccccctgcc cttctcctgg gacatcctgt
     1801 cccctcagtt catgtacggc tccagggcct tcatcaagca ccccgccgac atccccgact
     1861 actataagca gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg
     1921 gcggcgccgt gaccgtgacc caggacacct ccctggagga cggcaccctg atctacaagg
     1981 tgaagctcat cggcaccaac ttccctcctg acggccccgt aatgcagaag aagacaatgg
     2041 gctgggaagc gtccaccgag cggttgtacc ccgaggacgg cgtgctgaag ggcgacatta
     2101 agatggccct gcgcctgaag gacggcggcc gctacctggc ggacttcaag accacctaca
     2161 aggccaagaa gcccgtgcag atgcccggcg cctacaacgt cgaccgcaag ttggacatca
     2221 cctcccacaa cgaggactac accgtggtgg aacagtacga acgctccgag ggccgccact
     2281 ccaccggcgg catggacgag ccgtacaaat gagcggccgt gggccgccct caggtgcagg
     2341 ctgcctatca gaaggtggtg gctggtgtgg ccaatgccct ggctcacaaa taccactgag
     2401 atcgatcttt ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg
     2461 acttctggct aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt
     2521 ctctcactcg gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg
     2581 gtttagagtt tggcaacata tgcccatatg taactagcat aaccccttgg ggcctctaaa
     2641 cgggtcttga ggggtttttt gctgaaagca tgcggaggaa attctccttg aagtttccct
     2701 ggtgttcaaa gtaaaggagt ttgcaccaga cgcacctctg ttcactggtc cggcgtatta
     2761 aaacacgata cattgttatt agtacattta ttaagcgcta gattctgtgc gttgttgatt
     2821 tacagacaat tgttgtacgt attttaataa ttcattaaat ttataatctt tagggtggta
     2881 tgttagagcg aaaatcaaat gattttcagc gtctttatat ctgaatttaa atattaaatc
     2941 ctcaatagat ttgtaaaata ggtttcgatt agtttcaaac aagggttgtt tttccgaacc
     3001 gatggctgga ctatctaatg gattttcgct caacgccaca aaacttgcca aatcttgtag
     3061 cagcaatcta gctttgtcga tattcgtttg tgttttgttt tgtaataaag gttcgacgtc
     3121 gttcaaaata ttatgcgctt ttgtatttct ttcatcactg tcgttagtat acaattgact
     3181 cgacgtaaac acgttaaata gagcttggac atatttaaca tcgggcgtgt tagctttatt
     3241 aggccgatta tcgtcgtcgt cccaaccctc gtcgttagaa gttgcttccg aagacgattt
     3301 tgccatagcc acacgacgcc tattaattgt gtcggctaac acgtccgcga tcaaatttgt
     3361 agttgagctt tttggaattg cgatcgcata acttcgtata gcatacatta tacgaagtta
     3421 taagctcgga acgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     3481 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3541 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     3601 ccgccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     3661 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     3721 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     3781 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     3841 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     3901 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     3961 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     4021 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     4081 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     4141 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgtt agaaaaactc
     4201 atcgagcatc aagtgaaact gcaatttatt catatcagga ttatcaatac catatttttg
     4261 aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag
     4321 atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc
     4381 ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga
     4441 gaatggcaaa agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc
     4501 gtcatcaaaa tcactcgcac caaccaaacc gttattcatt cgtgattgcg cctgagcgag
     4561 acgaaatacg cgatcgccgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg
     4621 caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac
     4681 ctggaatgct gttttccctg ggatcgcagt ggtgagtaac catgcatcat caggagtacg
     4741 gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta gcctgaccat
     4801 ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc
     4861 atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc
     4921 ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc tcgagcaaga
     4981 cgtttcccgt tgaatatggc tcatactctt cctttttcaa tattattgaa gcatttatca
     5041 gggttattgt ctcatgtccg cgcgtttcct gcatctttta atcaaatccc aagatgtgta
     5101 taaacgcgcc ggtatgtata ggaagaggtt tatactaaac tgttacattg caaacgtggt
     5161 ttcgtgtgcc aagtgtgaaa accgatgttt aatcaaggct ctgacgcatt tctacaacca
     5221 cgactccaag tgtgtgggtg aagtcatgca tcttttaatc aaatcccaag atgtgtataa
     5281 accaccaaac tgccaaaaaa tgaaaactgt cgacaagctc tgtccgtttg ctggcaactg
     5341 caagggtctc aatcctattt gtaattattg aataataaaa caattataaa tgtcaaattt
     5401 gttttttatt aacgatacaa accaaacgca acaagaacat ttgtagtatt atctataatt
     5461 gaaaacgcgt agttataatc gctgaggtaa tatttaaaat cattttcaaa tgattcacag
     5521 ttaatttgcg acaatataat tttattttca cataaactag acgccttgtc gtcttcttct
     5581 tcgtattcct tctctttttc atttttctct tcataaaaat taacatagtt attatcgtat
     5641 ccatatatgt atctatcgta tagagtaaat tttttgttgt cataaatata tatgtctttt
     5701 ttaatggggt gtatagtacc gctgcgcata gtttttctgt aatttacaac agtgctattt
     5761 tctggtagtt cttcggagtg tgttgcttta attattaaat ttatataatc aatgaatttg
     5821 ggatcgtcgg ttttatacaa tatgttgccg gcatagtacg cagcttcttc tagttcaatt
     5881 acaccatttt ttagcagcac cggattaaca taactttcca aaatgttgta cgaaccgtta
     5941 aacaaaaaca gttcacctcc cttttctata ctattgtctg cgagcagttg tttgttgtta
     6001 aaaataacag ccattgtaat gagacgcaca aactaatatc acaaactgga aatgtctatc
     6061 aatatatagt tgctgatggc cggcctatta atagtaatca attacggggt cattagttca
     6121 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
     6181 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
     6241 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
     6301 acatcaagtg tatcatatgc caagtccgcc ccctattgac gtcaatgacg gtaaatggcc
     6361 cgcctggcat tatgcccagt acatgacctt acgggacttt cctacttggc agtacatcta
     6421 cgtattagtc atcgctatta ccatgctgat gcggttttgg cagtacacca atgggcgtgg
     6481 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
     6541 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aataaccccg ccccgttgac
     6601 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagacgtc gtttagtgaa
     6661 ccgtcagatc actagatgct ttattgcggt agtttatcac agttaaattg ctaacgccag
     6721 tctcgaactt aacgtgcaga agttggtcgt gaggcactgg gcaggtaagt atcgggccct
     6781 ttgtgcgggg ggagcggctc ggggctgtcc gcggggggac ggctgccttc gggggg
//
