LOCUS       sequence-273573-        6734 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Booster-ROCK, a Red-Shifted Genetically Encoded FRET Biosensor for 
            ROCK activity..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6734)
  AUTHORS   Watabe T, Terai K, Sumiyama K, Matsuda M
  TITLE     Booster, a Red-Shifted Genetically Encoded Forster Resonance Energy 
            Transfer (FRET) Biosensor Compatible with Cyan Fluorescent 
            Protein/Yellow Fluorescent Protein-Based FRET Biosensors and Blue 
            Light-Responsive Optogenetic Tools.
  JOURNAL   ACS Sens. 2020 Feb 26. doi: 10.1021/acssensors.9b01941.
  PUBMED    32101394
REFERENCE   2  (bases 1 to 6734)
  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: "ACS Sens. 
            2020 Feb 26. doi: 10.1021/acssensors.9b01941."
COMMENT     Sequence Label: pCAGGS-4591NES
FEATURES             Location/Qualifiers
     source          1..6734
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          610..1630
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1550..1572
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1638..1657
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1687..1696
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1693..2388
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     CDS             2986..3639
                     /codon_start=1
                     /product="monomeric Kusabira-Orange 2 fluorescent protein"
                     /label=mKO2
                     /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV
                     TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA
                     SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY
                     LKLEGGGNHKCQFKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAHS"
     CDS             3706..3732
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     primer_bind     complement(3748..3767)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3813..3868
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3867..3886)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4229..4245)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4229..4245)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4242..4264)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4253..4269
                     /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(4277..4283,4284..4301,4302..4307))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4322..4343
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4402..4597
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4448..4583
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4603..4737
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4640..4659)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4694..4713
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4805..4822)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4976..5564)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5056..5075)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5735..6526,6527..6595))
                     /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     6358..6377
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6596..6700)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact ctccccatct
      421 cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt tgtgcagcga
      481 tgggggcggg ggggggcgag gcggagaggt gcggcggcag ccaatcagag cggcgcgctc
      541 cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg
      601 cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg
      661 cgccgcccgc cccggctctg actgaccgcg ttactcccac aggtgagcgg gcgggacggc
      721 ccttctcctc cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg
      781 ctgcgtgaaa gccttgaggg gctccgggag ggccctttgt gcggggggag cggctcgggg
      841 ggtgcgtgcg tgtgtgtgtg cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc
      901 tgtgagcgct gcgggcgcgg cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc
      961 gcggccgggg gcggtgcccc gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg
     1021 cggggtgtgt gcgtgggggg gtgagcaggg ggtgtgggcg cgtcggtcgg gctgcaaccc
     1081 cccctgcacc cccctccccg agttgctgag cacggcccgg cttcgggtgc ggggctccgt
     1141 acggggcgtg gcgcggggct cgccgtgccg ggcggggggt ggcggcaggt gggggtgccg
     1201 ggcggggcgg ggccgcctcg ggccggggag ggctcggggg aggggcgcgg cggcccccgg
     1261 agcgccggcg gctgtcgagg cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc
     1321 gagagggcgc agggacttcc tttgtcccaa atctgtgcgg agccgaaatc tgggaggcgc
     1381 cgccgcaccc cctctagcgg gcgcggggcg aagcggtgcg gcgccggcag gaaggaaatg
     1441 ggcggggagg gccttcgtgc gtcgccgcgc cgccgtcccc ttctccctct ccagcctcgg
     1501 ggctgtccgc ggggggacgg ctgccttcgg gggggacggg gcagggcggg gttcggcttc
     1561 tggcgtgtga ccggcggctc tagctagagc ctctgctaac catgttcatg ccttcttctt
     1621 tttcctacag ctcctgggca acgtgctggt tattgtgctg tctcatcatt ttggcaagaa
     1681 ttcgccgcca ccatggtgag cgagctgatt aaggagaaca tgcacatgaa gctgtacatg
     1741 gagggcaccg tgaacaacca ccacttcaag tgcacatccg agggcgaagg caagccctac
     1801 gagggcaccc agaccatgag aatcaaggcg gtcgagggcg gccctctccc cttcgccttc
     1861 gacatcctgg ctaccagctt catgtacggc agcaaaacct tcatcaacca cacccagggc
     1921 atccccgact tctttaagca gtccttcccc gagggcttca catgggagag agtcaccaca
     1981 tacgaagacg ggggcgtgct gaccgctacc caggacacca gcctccagga cggctgcctc
     2041 atctacaacg tcaagatcag aggggtgaac ttcccatcca acggccctgt gatgcagaag
     2101 aaaacactcg gctgggaggc ctccaccgag accctgtacc ccgctgacgg cggcctggaa
     2161 ggcagagccg acatggccct gaagctcgtg ggcgggggcc acctgatctg caacttgaag
     2221 accacataca gatccaagaa acccgctaag aacctcaaga tgcccggcgt ctactatgtg
     2281 gacagaagac tggaaagaat caaggaggcc gacaaagaga cctacgtcga gcagcacgag
     2341 gtggctgtgg ccagatactg cgacctccct agcaaactgg ggcacagact cgagaagttt
     2401 tctcaagaac agatcggcga aaacattgtg tgcagggtca tttgtaccac gggtcaaatt
     2461 cccatccgag atttgtcagc tgatatttca caagtgctta aggaaaaacg atccataaag
     2521 aaagtttgga catttggtag aaacccagcc tgtgactatc atttaggaaa catttcaaga
     2581 ctgtcaaata agcatttcca aatactacta ggagaagacg gtaacctttt attgaatgac
     2641 atttccacta atgggacctg gttaaatggg caaaaagtcg agaagaacag caatcagtta
     2701 ctgtcccaag gtgatgaaat aaccgttggt gtaggcgtgg aatcagatat tttatctctg
     2761 gtcattttca taaacgacaa atttaagcag tgcctggagc agaacaaagt tgatcgcggt
     2821 accagtgctg gtggtagtgc tggtggtagt gctggtggta gtgctggtgg tagtgctggt
     2881 ggttccggca gtgctggtgg tagtgctggt ggtagtacca gtgctggtgg tagtgctggt
     2941 ggtagtgctg gtggtagtgc tggtggtagt gctggtggtt ccggaatggt gagtgtgatt
     3001 aaaccagaga tgaagatgag gtactacatg gacggctccg tcaatgggca tgagttcaca
     3061 attgaaggtg aaggcacagg cagaccttac gagggacatc aagagatgac actacgcgtc
     3121 acaatggccg agggcgggcc aatgcctttc gcgtttgact tagtgtcaca cgtgttctgt
     3181 tacggccaca gagtatttac taaatatcca gaagagatac cagactattt caaacaagca
     3241 tttcctgaag gcctgtcatg ggaaaggtcg ttggagttcg aagatggtgg gtccgcttca
     3301 gtcagtgcgc atataagcct tagaggaaac accttctacc acaaatccaa atttactggg
     3361 gttaactttc ctgccgatgg tcctatcatg caaaaccaaa gtgttgattg ggagccatca
     3421 accgagaaaa ttactgccag cgacggagtt ctgaagggtg atgttacgat gtacctaaaa
     3481 cttgaaggag gcggcaatca caaatgccaa ttcaagacta cttacaaggc ggcaaaagag
     3541 attcttgaaa tgccaggaga ccattacatc ggccatcgcc tcgtcaggaa aaccgaaggc
     3601 aacattactg agcaggtaga agatgcagta gctcattccg cggccgcaaa gagaagaaac
     3661 agaagaaaga ccctggtaga tctgcctctg gattctagac tgcagctgcc tcctctggaa
     3721 cgcctgactc tggattaagt cgacggatct ttttccctct gccaaaaatt atggggacat
     3781 catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt tcattgcaat
     3841 agtgtgttgg aattttttgt gtctctcact cggaaggaca tatgggaggg caaatcattt
     3901 aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgcccata tgctggctgc
     3961 catgaacaaa ggttggctat aaagaggtca tcagtatatg aaacagcccc ctgctgtcca
     4021 ttccttattc catagaaaag ccttgacttg aggttagatt ttttttatat tttgttttgt
     4081 gttatttttt tctttaacat ccctaaaatt ttccttacat gttttactag ccagattttt
     4141 cctcctctcc tgactactcc cagtcatagc tgtccctctt ctcttatgga gatccctcga
     4201 cctgcagccc aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     4261 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     4321 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     4381 acctgtcgtg ccagcggatc cgcatctcaa ttagtcagca accatagtcc cgcccctaac
     4441 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     4501 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     4561 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctaacttgtt tattgcagct
     4621 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     4681 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatccgc
     4741 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     4801 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4861 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4921 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4981 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     5041 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     5101 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     5161 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5221 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5281 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5341 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5401 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5461 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5521 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5581 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5641 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5701 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5761 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5821 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     5881 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5941 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     6001 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     6061 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     6121 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6181 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6241 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6301 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6361 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6421 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     6481 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6541 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     6601 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6661 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6721 cacctgggtc gatc
//
