LOCUS       sequence-204833-        7939 bp    DNA     circular SYN 14-APR-2025
DEFINITION  A S6 Kinase biosensor based on fluorescence resonance energy 
            transfer (FRET) and those based on bioluminescence energy transfer 
            (BRET).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7939)
  AUTHORS   Komatsu N, Terai K, Imanishi A, Kamioka Y, Sumiyama K, Jin T, Okada 
            Y, Nagai T, Matsuda M
  TITLE     A platform of BRET-FRET hybrid biosensors for optogenetics, chemical
            screening, and in vivo imaging.
  JOURNAL   Sci Rep. 2018 Jun 12;8(1):8984. doi: 10.1038/s41598-018-27174-x.
  PUBMED    29895862
REFERENCE   2  (bases 1 to 7939)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Rep."; 
            date: "2018-06-12"; volume: "8(1):8984. doi"; pages: " 
            10.1038/s41598-018-27174-x"
COMMENT     Sequence Label: pCAGGS-6015nes
FEATURES             Location/Qualifiers
     source          1..7939
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        8..387
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..665
                     /label=chicken beta-actin promoter
     intron          666..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1598..1620
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     CDS             join(1735..1737,1738..1740,1741..2448)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLLCTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALFKDPNEKRDHMVLL
                     EFLTAAGITEGMNELY"
     CDS             4921..4947
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     primer_bind     complement(4963..4982)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    5028..5083
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5082..5101)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5442..5458)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5442..5458)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5455..5477)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5466..5482
                     /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(5490..5496,5497..5514,5515..5520))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5535..5556
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5615..5810
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5661..5796
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5816..5950
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5853..5872)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5907..5926
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6018..6035)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6189..6777)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6269..6288)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6948..7739,7740..7808))
                     /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     7571..7590
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7809..7913)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgatcgac attgattatt gactagttat taatagtaat caattacggg gtcattagtt
       61 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
      121 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
      181 atagggactt tccattgacg tcaatgggtg gactatttac ggtaaactgc ccacttggca
      241 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
      301 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
      361 tacgtattag tcatcgctat taccatgggt cgaggtgagc cccacgttct gcttcactct
      421 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
      481 tgcagcgatg ggggcggggg gggggggggc gcgcgccagg cggggcgggg cgggggcgag
      541 ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa tcagagcggc gcgctccgaa
      601 agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga agcgcgcggc
      661 gggcgggagt cgctgcgttg ccttcgcccc gtgccccgct ccgcgccgcc tcgcgccgcc
      721 cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc
      781 ctccgggctg taattagcgc ttggtttaat gacggctcgt ttcttttctg tggctgcgtg
      841 aaagccttaa agggctccgg gagggccctt tgtgcggggg ggagcggctc ggggggtgcg
      901 tgcgtgtgtg tgtgcgtggg gagcgccgcg tgcggcccgc gctgcccggc ggctgtgagc
      961 gctgcgggcg cggcgcgggg ctttgtgcgc tccgcgtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg ggggggctgc gaggggaaca aaggctgcgt gcggggtgtg
     1081 tgcgtggggg ggtgagcagg gggtgtgggc gcggcggtcg ggctgtaacc cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtgcggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg agggctcggg ggaggggcgc ggcggccccg gagcgccggc
     1321 ggctgtcgag gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg cgagagggcg
     1381 cagggacttc ctttgtccca aatctggcgg agccgaaatc tgggaggcgc cgccgcaccc
     1441 cctctagcgg gcgcgggcga agcggtgcgg cgccggcagg aaggaaatgg gcggggaggg
     1501 ccttcgtgcg tcgccgcgcc gccgtcccct tctccatctc cagcctcggg gctgccgcag
     1561 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     1621 ggcggctcta gctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggttg ttgtgctgtc tcatcatttt ggcaagaatt cggcatggtg
     1741 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     1801 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     1861 ctgaccctga agcttctatg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     1921 accaccctgg gctacggcct gcagtgcttc gcccgctacc ccgaccacat gaagcagcac
     1981 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     2041 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     2101 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     2161 gagtacaact acaacagcca caacgtctat atcaccgccg acaagcagaa gaacggcatc
     2221 aaggccaact tcaagatccg ccacaacatc gaggacggcg gcgtgcagct cgccgaccac
     2281 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     2341 agctaccagt ccgccctgtt caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     2401 gagttcctga ccgccgccgg gatcactgag ggcatgaacg agctgtacct cgagaagttt
     2461 tctcaagaac agatcggcga aaacattgtg tgcagggtca tttgtaccac gggtcaaatt
     2521 cccatccgag atttgtcagc tgatatttca caagtgctta aggaaaaacg atccataaag
     2581 aaagtttgga catttggtag aaacccagcc tgtgactatc atttaggaaa catttcaaga
     2641 ctgtcaaata agcatttcca aatactacta ggagaagacg gtaacctttt attgaatgac
     2701 atttccacta atgggacctg gttaaatggg caaaaagtcg agaagaacag caatcagtta
     2761 ctgtcccaag gtgatgaaat aaccgttggt gtaggcgtgg aatcagatat tttatctctg
     2821 gtcattttca taaacgacaa atttaagcag tgcctggagc agaacaaagt tgatcgcggt
     2881 accagtgctg gtggtagtgc tggtggtagt gctggtggta gtgctggtgg tagtgctggt
     2941 ggttccggca gtgctggtgg tagtgctggt ggtagtacca gtgctggtgg tagtgctggt
     3001 ggtagtgctg gtggtagtgc tggtggtagt gctggtggtt ccggcagtgc tggtggtagt
     3061 gctggtggta gtaccagtgc tggtggtagt gctggtggta gtgctggtgg tagtgctggt
     3121 ggtagtgctg gtggttccgg cagtgctggt ggtagtgctg gtggtagtac cagtgctggt
     3181 ggtagtgctg gtggtagtgc tggtggtagt gctggtggta gtgctggtgg ttccggaaac
     3241 aggcgaatca gaacacttac ggagcccgac gttgatttta atggcggccg catggtgagc
     3301 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgtc
     3361 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
     3421 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
     3481 accctgtcct ggggcgtgca gtgcttcgcc cgctaccccg accacatgaa gcagcacgac
     3541 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
     3601 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
     3661 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
     3721 tacaactact tcagcgggaa cgtctatatc accgccgaca agcagaagaa cggcatcaag
     3781 gccaacttca agatccgcca caacatcgag gacggcggcg tgcagctcgc cgaccactac
     3841 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     3901 acccagtccg ccttaagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     3961 ttcttgaccg ccgccggggg taccaaggtg tacgaccccg agcagaggaa gaggatgatc
     4021 accggccccc agtggtgggc caggtgcaag cagatgaacg tgctggacag cttcatcaac
     4081 tactacgaca gcgagaagca cgccgagaac gccgtgatct tcctgcacgg caacgccact
     4141 agcagctacc tgtggaggca cgtggtgccc cacatcgagc ccgtggccag gtgcatcatc
     4201 cccgatctga tcggcatggg caagagcggc aagagcggca acggcagcta caggctgctg
     4261 gaccactaca agtacctgac cgcctggttc gagcttctga acctgcccaa gaagatcatc
     4321 ttcgtgggcc acgactgggg cgccgccctg gccttccact acgcctacga gcaccaggac
     4381 aggatcaagg ccatcgtgca catggagagc gtggtggacg tgatcgagag ctgggacgag
     4441 tggccagaca tcgaggagga catcgccctg atcaagagcg aggagggcga gaagatggtg
     4501 ctggagaaca acttcttcgt ggagaccgtg ctgcccagca agatcatgag aaagctggag
     4561 cccgaggagt tcgccgccta cctggagccc ttcaaggaga agggcgaggt gagaagaccc
     4621 accctgagct ggcccagaga gatccccctg gtgaagggcg gcaagcccga cgtggtgcag
     4681 atcgtgagaa actacaacgc ctacctgaga gccagcgacg acctgcccaa gctgttcatc
     4741 gagggcgacc ccggcttctt cagcaacgcc atcgtggagg gcgccaagaa gttccccaac
     4801 accgagttcg tgaaggtgaa gggcctgcac ttcctccagg aggacgcccc cgacgagatg
     4861 ggcaagtaca tcaagagctt cgtggagaga gtgctgaaga acgagcagtc tagactgcag
     4921 ctgcctcctc tggaacgcct gactctggat taagtcgacg gatctttttc cctctgccaa
     4981 aaattatggg gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt
     5041 tattttcatt gcaatagtgt gttggaattt tttgtgtctc tcactcggaa ggacatatgg
     5101 gagggcaaat catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc
     5161 catatgctgg ctgccatgaa caaaggtggc tataaagagg tcatcagtat atgaaacagc
     5221 cccctgctgt ccattcctta ttccatagaa aagccttgac ttgaggttag attttttttt
     5281 atattttgtt tgtgttattt ttttctttaa catccctaaa attttcctta catgttttac
     5341 tagccagatt tttcctcctc tcctgactac tcccagtcat agctgtccct cttctcttat
     5401 gaagatccct cgacctgcag cccaagcttg gcgtaatcat ggtcatagct gtttcctgtg
     5461 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     5521 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     5581 ttccagtcgg gaaacctgtc gtgccagcgg atccgcatct caattagtca gcaaccatag
     5641 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     5701 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     5761 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctaactt
     5821 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     5881 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     5941 tgtctggatc cgctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     6001 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     6061 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     6121 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     6181 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     6241 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     6301 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     6361 tcgggaagcg tggcgctttc tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc
     6421 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     6481 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     6541 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     6601 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     6661 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     6721 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     6781 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     6841 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     6901 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     6961 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     7021 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     7081 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     7141 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     7201 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     7261 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     7321 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     7381 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     7441 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     7501 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     7561 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     7621 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     7681 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     7741 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     7801 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     7861 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     7921 ccccgaaaag tgccacctg
//
