LOCUS       sequence-353480-        6511 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FRET biosensor. The highly sensitive biosensor has a mutated SH2 
            domain, ECFP and YPet.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6511)
  AUTHORS   Ouyang M, Wan R, Qin Q, Peng Q, Wang P, Wu J, Allen M, Shi Y, Laub 
            S, Deng L, Lu S, Wang Y
  TITLE     Sensitive FRET Biosensor Reveals Fyn Kinase Regulation by 
            Submembrane Localization.
  JOURNAL   ACS Sens. 2019 Jan 25;4(1):76-86. doi: 10.1021/acssensors.8b00896. 
            Epub 2019 Jan 9.
  PUBMED    30588803
REFERENCE   2  (bases 1 to 6511)
  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"; doi: 
            "10.1021/acssensors.8b00896"; journalName: "ACS Sens"; date: 
            "2019-01-25- 25"; volume: "4"; issue: "1"; pages: "76-86"
COMMENT     Sequence Label: Cytosolic Fyn FRET Biosensor
FEATURES             Location/Qualifiers
     source          1..6511
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        3..382
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        384..659
                     /label=chicken beta-actin promoter
     intron          660..1680
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1600..1622
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1688..1707
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             join(1736..1738,1739..1741,1742..2254)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     AHFKIRHNIE"
     primer_bind     complement(1781..1802)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2042..2061)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2389..2410
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2804..3514
                     /codon_start=1
                     /product="yellow fluorescent protein optimized for FRET 
                     with CyPet (Nguyen and Daugherty, 2005)"
                     /label=YPet
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKL
                     LCTTGKLPVPWPTLVTTLGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKAN
                     FKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALFKDPNEKRDHMVLLEF
                     LTAAGITEGMNELYK"
     primer_bind     complement(3527..3546)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3592..3647
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3646..3665)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4008..4024)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4008..4024)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4021..4043)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4032..4048
                     /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(4056..4062,4063..4080,4081..4086))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4101..4122
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4181..4376
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4227..4362
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4382..4516
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4419..4438)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4473..4492
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4584..4601)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4755..5343)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4835..4854)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5514..6305,6306..6374))
                     /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     6137..6156
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6375..6479)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
       61 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
      121 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
      181 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
      241 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
      301 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
      361 attagtcatc gctattacca tggtcgaggt gagccccacg ttctgcttca ctctccccat
      421 ctcccccccc tccccacccc caattttgta tttatttatt ttttaattat tttgtgcagc
      481 gatgggggcg gggggggggg gggcgcgcgc caggcggggc ggggcggggc gaggggcggg
      541 gcggggcgag gcggagaggt gcggcggcag ccaatcagag cggcgcgctc cgaaagtttc
      601 cttttatggc gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg cggcgggcgg
      661 gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc
      721 cccggctctg actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc
      781 cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa
      841 gccttgaggg gctccgggag ggccctttgt gcggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc tgtgagcgct
      961 gcgggcgcgg cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc gcggccgggg
     1021 gcggtgcccc gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg cggggtgtgt
     1081 gcgtgggggg gtgagcaggg ggtgtgggcg cgtcggtcgg gctgcaaccc cccctgcacc
     1141 cccctccccg agttgctgag cacggcccgg cttcgggtgc ggggctccgt acggggcgtg
     1201 gcgcggggct cgccgtgccg ggcggggggt ggcggcaggt gggggtgccg ggcggggcgg
     1261 ggccgcctcg ggccggggag ggctcggggg aggggcgcgg cggcccccgg agcgccggcg
     1321 gctgtcgagg cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc gagagggcgc
     1381 agggacttcc tttgtcccaa atctgtgcgg agccgaaatc tgggaggcgc cgccgcaccc
     1441 cctctagcgg gcgcggggcg aagcggtgcg gcgccggcag gaaggaaatg ggcggggagg
     1501 gccttcgtgc gtcgccgcgc cgccgtcccc ttctccctct ccagcctcgg ggctgtccgc
     1561 ggggggacgg ctgccttcgg gggggacggg gcagggcggg gttcggcttc tggcgtgtga
     1621 ccggcggctc tagctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag
     1681 ctcctgggca acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attctatggt
     1741 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     1801 cgtaaacggc cacaggttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     1861 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     1921 gaccaccctg acctggggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     1981 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgtacca tcttcttcaa
     2041 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     2101 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     2161 ggagtacaac tacatcagcc acaacgtcta tatcaccgcc gacaagcaga agaacggcat
     2221 caaggcccac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     2281 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     2341 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     2401 ggagttcgtg accgccgccc gcatgcattg gtattttggg aagatcactc gtcgggagtc
     2461 cgagcggctg ctgctcaacc ccgaaaaccc ccggggaacc ttcttggtcc gggagagcga
     2521 gacgacaaaa ggtgcctatg ccctctccgt ttctgacttt gacaacgcca aggggctcaa
     2581 tgtgaagcac tacaagatcc gcaagctgga cagcggcggc ttctacatca cctcacgcac
     2641 acagttcagc agcctgcagc agctggtggc ctactactcc aaacatgctg atggcttgtg
     2701 ccaccgcctg actaacgtct gtgggtctac atctggatct gggaagccgg gttctggtga
     2761 gggttctgag aagatcgagg gcacctacgg cgtcgtggag ctctctaaag gtgaagaatt
     2821 attcactggt gttgtcccaa ttttggttga attagatggt gatgttaatg gtcacaaatt
     2881 ttctgtctcc ggtgaaggtg aaggtgatgc tacgtacggt aaattgacct taaaattact
     2941 ctgtactact ggtaaattgc cagttccatg gccaacctta gtcactactt taggttatgg
     3001 tgttcaatgt tttgctagat acccagatca tatgaaacaa catgactttt tcaagtctgc
     3061 catgccagaa ggttatgttc aagaaagaac tatttttttc aaagatgacg gtaactacaa
     3121 gaccagagct gaagtcaagt ttgaaggtga taccttagtt aatagaatcg aattaaaagg
     3181 tattgatttt aaagaagatg gtaacatttt aggtcacaaa ttggaataca actataactc
     3241 tcacaatgtt tacatcactg ctgacaaaca aaagaatggt atcaaagcta acttcaaaat
     3301 tagacacaac attgaagatg gtggtgttca attagctgac cattatcaac aaaatactcc
     3361 aattggtgat ggtccagtct tgttaccaga caaccattac ttatcctatc aatctgcctt
     3421 attcaaagat ccaaacgaaa agagagacca catggtcttg ttagaatttt tgactgctgc
     3481 tggtattacc gagggtatga atgaattgta caaataagtc gacggatctt tttccctctg
     3541 ccaaaaatta tggggacatc atgaagcccc ttgagcatct gacttctggc taataaagga
     3601 aatttatttt cattgcaata gtgtgttgga attttttgtg tctctcactc ggaaggacat
     3661 atgggagggc aaatcattta aaacatcaga atgagtattt ggtttagagt ttggcaacat
     3721 atgcccatat gctggctgcc atgaacaaag gttggctata aagaggtcat cagtatatga
     3781 aacagccccc tgctgtccat tccttattcc atagaaaagc cttgacttga ggttagattt
     3841 tttttatatt ttgttttgtg ttattttttt ctttaacatc cctaaaattt tccttacatg
     3901 ttttactagc cagatttttc ctcctctcct gactactccc agtcatagct gtccctcttc
     3961 tcttatggag atccctcgac ctgcagccca agcttggcgt aatcatggtc atagctgttt
     4021 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     4081 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     4141 cccgctttcc agtcgggaaa cctgtcgtgc cagcggatcc gcatctcaat tagtcagcaa
     4201 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     4261 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
     4321 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     4381 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     4441 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4501 ttatcatgtc tggatccgct gcattaatga atcggccaac gcgcggggag aggcggtttg
     4561 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     4621 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     4681 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4741 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4801 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4861 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4921 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4981 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     5041 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     5101 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     5161 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     5221 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     5281 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     5341 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     5401 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     5461 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     5521 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     5581 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     5641 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     5701 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     5761 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     5821 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     5881 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     5941 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     6001 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     6061 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     6121 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     6181 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     6241 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     6301 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     6361 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     6421 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     6481 acatttcccc gaaaagtgcc acctgggtcg a
//
