LOCUS       sequence-111419-        6166 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FRET biosensor Destination vector for E. coli expression. Encodes a 
            FRET pair for fusion with ligand binding domain of interest..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6166)
  AUTHORS   Jones AM, Danielson JA, Manojkumar SN, Lanquar V, Grossmann G, 
            Frommer WB
  TITLE     Abscisic acid dynamics in roots detected with genetically encoded 
            FRET sensors.
  JOURNAL   eLife. 2014 Apr 15;3:e01741. doi: 10.7554/eLife.01741.
  PUBMED    24737862
REFERENCE   2  (bases 1 to 6166)
  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: "eLife."; 
            date: "2014-04-15"; volume: "3:e01741. doi"; pages: " 
            10.7554/eLife.01741"
COMMENT     Sequence Label: pFLIP42
FEATURES             Location/Qualifiers
     source          1..6166
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     19..38
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        19..37
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             69..91
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             111..128
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             132..164
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(168..176,177..191)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(207..209,210..920)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     protein_bind    933..1057
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1082..1087,1088..1105,1106..1112)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     primer_bind     complement(1232..1251)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2225..2530
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2409..2428
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2571..2695)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             join(2709..2711,2712..3422)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             3423..3452
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3511..3529)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3525..3572
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3669..4124
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3756..3775)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3966..3987
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4205..4309
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4310..4378,4379..5170)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCDTLLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDESDTTMPVAMPTTLRKLLTGELLTLASRQQLID*MEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKRSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5341..5929
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5830..5849
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6083..6100
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 atctcgatcc cgcgaaatta atacgactca ctatagggag accacaacgg tttccctcta
       61 gaaataattt tgtttaactt taagaaggag atatacatat gcggggttct catcatcatc
      121 atcatcatgg tatggctagc atgactggtg gacagcaaat gggtcgggat ctgtacgacg
      181 atgacgataa ggatccgggc ctcgaggtga gcaagggcga ggagctgttc accggggtgg
      241 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
      301 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
      361 agctgcccgt gccctggccc accctcgtga ccaccttcgg ctacggcctg atgtgcttcg
      421 cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
      481 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
      541 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
      601 aggacggcaa catcctgggg cacaagcttg agtacaacta caacagccac aacgtctata
      661 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg
      721 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
      781 ccgtgctgct gcccgacaac cactacctga gctaccagtc cgccctgagc aaagacccca
      841 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
      901 gcatggacga gctgtacaag ggtaccgata tcacaagttt gtacaaaaaa gctgaacgag
      961 aaacgtaaaa tgatataaat atcaatatat taaattagat tttgcataaa aaacagacta
     1021 cataatactg taaaacacaa catatccagt cactatggcg gccgcattag gcaccccagg
     1081 ctttacactt tatgcttccg gctcgtataa tgtgtggatt ttgagttagg atccgtcgag
     1141 attttcagga gctaaggaag ctaaaatgga gaaaaaaatc actggatata ccaccgttga
     1201 tatatcccaa tggcatcgta aagaacattt tgaggcattt cagtcagttg ctcaatgtac
     1261 ctataaccag accgttcagc tggatattac ggccttttta aagaccgtaa agaaaaataa
     1321 gcacaagttt tatccggcct ttattcacat tcttgcccgc ctgatgaatg ctcatccgga
     1381 attccgtatg gcaatgaaag acggtgagct ggtgatatgg gatagtgttc acccttgtta
     1441 caccgttttc catgagcaaa ctgaaacgtt ttcatcgctc tggagtgaat accacgacga
     1501 tttccggcag tttctacaca tatattcgca agatgtggcg tgttacggtg aaaacctggc
     1561 ctatttccct aaagggttta ttgagaatat gtttttcgtc tcagccaatc cctgggtgag
     1621 tttcaccagt tttgatttaa acgtggccaa tatggacaac ttcttcgccc ccgttttcac
     1681 catgggcaaa tattatacgc aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca
     1741 tcatgccgtt tgtgatgggc ttccatgtcg gcagaatgct taatgaatta cacagtactg
     1801 cgatgagtgg cagggcgggg cgtaaacgcg tggatccggc ttactaaaag ccagataaca
     1861 gtatgcgtat ttgcgcgctg atttttgcgg tataagaata tatactgata tgtatacccg
     1921 aagtatgtca aaaagaggta tgctatgaag cagcgtatta cagtgacagt tgacagcgac
     1981 agctatcagt tgctcaaggc atatatgatg tcaatatctc cggtctggta agcacaacca
     2041 tgcagaatga agcccgtcgt ctgcgtgccg aacgctggaa agcggaaaat caggaaggga
     2101 tggctgaggt cgcccggttt attgaaatga acggctcttt tgctgacgag aacaggggct
     2161 ggtgaaatgc agtttaaggt ttacacctat aaaacttttg ctgacgagaa caggggctgg
     2221 tgaaatgcag tttaaggttt acacctataa aagagagagc cgttatcgtc tgtttgtgga
     2281 tgtacagagt gatattattg acacgcccgg gcgacggatg gtgatccccc tggccagtgc
     2341 acgtctgctg tcagataaag tctcccgtga actttacccg gtggtgcata tcggggatga
     2401 aagctggcgc atgatgacca ccgatatggc cagtgtgccg gtctccgtta tcggggaaga
     2461 agtggctgat ctcagccacc gcgaaaatga catcaaaaac gccattaacc tgatgttctg
     2521 gggaatataa atgtcaggct cccttataca cagccagtct gcaggtcgac catagtgact
     2581 ggatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc taatttaata
     2641 tattgatatt tatatcattt tacgtttctc gttcagcttt cttgtacaaa gtggtgatat
     2701 cgactagtgt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2761 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     2821 cctacggcaa gctgaccctg aagttcatct gcaccaccgg taagctgccc gtgccctggc
     2881 ccaccctcgt gaccaccctg acctggggcg tgcagtgctt cgcccgctac cccgaccaca
     2941 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     3001 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     3061 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     3121 ggcacaagct ggagtacaac gccatcagcg acaacgtcta tatcaccgcc gacaagcaga
     3181 agaacggcat caaggccaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     3241 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     3301 accactacct gagcacccag tccaagctga gcaaagaccc caacgagaag cgcgatcaca
     3361 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     3421 aggaacaaaa attgataagt gaggaagatt tataagggcc cgatccggct gctaacaaag
     3481 cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg
     3541 gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata tccggatctg
     3601 gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg
     3661 cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     3721 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     3781 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     3841 ccgatttaga gctttacggc acctcgaccg caaaaaactt gatttgggtg atggttcacg
     3901 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     3961 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatcgcgg tctattcttt
     4021 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     4081 aatatttaac gcgaatttta acaaaatatt aacgtttaca atttcgcctg atgcggtatt
     4141 ttctccttac gcatctgtgc ggtatttcac accgcataca ggtggcactt ttcggggaaa
     4201 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     4261 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     4321 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     4381 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     4441 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     4501 tccaatgatg agcactttta aagttctgct atgtgataca ctattatccc gtattgacgc
     4561 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     4621 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     4681 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     4741 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     4801 accggagctg aatgaagcca taccaaacga cgagagtgac accacgatgc ctgtagcaat
     4861 gccaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     4921 attaatagac tgaatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     4981 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     5041 tgcagcactg gggccagatg gtaagcgctc ccgtatcgta gttatctaca cgacggggag
     5101 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     5161 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     5221 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     5281 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     5341 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     5401 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     5461 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     5521 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     5581 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     5641 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     5701 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     5761 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     5821 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     5881 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     5941 cgcggccttt ttacggttcc tgggcttttg ctggcctttt gctcacatgt tctttcctgc
     6001 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     6061 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     6121 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagg
//
