LOCUS       sequence-111222-        6103 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 6103)
  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 6103)
  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: pFLIP35
FEATURES             Location/Qualifiers
     source          1..6103
                     /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"
     protein_bind    906..1030
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1055..1060,1061..1078,1079..1085)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     primer_bind     complement(1205..1224)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2198..2503
                     /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     2382..2401
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2544..2668)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             3360..3389
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3448..3466)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3462..3509
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3606..4061
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3693..3712)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3903..3924
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4142..4246
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4247..4315,4316..5107)
                     /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      5278..5866
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5767..5786
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6020..6037
                     /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 ctcgaggttt ctaaaggtga agaattgttt acgggcgtcg
      241 tcccgatcct cgtggaactc gacggggatg ttaacgggca taagttttcg gtcagcgggg
      301 aaggggaggg ggacgcgacg tatgggaagc tcactctcaa gctgatctgt acgacgggga
      361 aactcccggt cccgtggccg acgctggtca cgacgctggg atacgggctc caatgctttg
      421 cgaggtatcc ggaccacatg aaacagcatg actttttcaa atcggcgatg ccggagggat
      481 acgtgcagga acggacgatc tttttcaaag acgatgggaa ctataagacg cgggcggaag
      541 tcaagtttga aggggacacg ctcgtcaacc ggatcgaact caaggggatt gacttcaaag
      601 aggatgggaa catactcggc cataagctcg aatacaatta caactcgcat aacgtataca
      661 tcaccgcgga taagcaaaag aacgggatca aagccaattt caaaatccgg cataacatag
      721 aggatggggg ggtccaactg gcggatcact atcagcaaaa cacgccgata ggggatgggc
      781 cggtcctcct cccggataac cattacctct cgtaccaaag cgcgctctcg aaggacccga
      841 atgagaaacg ggaccacatg gttctcctgg agttcgtcac ggcggcgggc ataggtaccg
      901 atatcacaag tttgtacaaa aaagctgaac gagaaacgta aaatgatata aatatcaata
      961 tattaaatta gattttgcat aaaaaacaga ctacataata ctgtaaaaca caacatatcc
     1021 agtcactatg gcggccgcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     1081 taatgtgtgg attttgagtt aggatccgtc gagattttca ggagctaagg aagctaaaat
     1141 ggagaaaaaa atcactggat ataccaccgt tgatatatcc caatggcatc gtaaagaaca
     1201 ttttgaggca tttcagtcag ttgctcaatg tacctataac cagaccgttc agctggatat
     1261 tacggccttt ttaaagaccg taaagaaaaa taagcacaag ttttatccgg cctttattca
     1321 cattcttgcc cgcctgatga atgctcatcc ggaattccgt atggcaatga aagacggtga
     1381 gctggtgata tgggatagtg ttcacccttg ttacaccgtt ttccatgagc aaactgaaac
     1441 gttttcatcg ctctggagtg aataccacga cgatttccgg cagtttctac acatatattc
     1501 gcaagatgtg gcgtgttacg gtgaaaacct ggcctatttc cctaaagggt ttattgagaa
     1561 tatgtttttc gtctcagcca atccctgggt gagtttcacc agttttgatt taaacgtggc
     1621 caatatggac aacttcttcg cccccgtttt caccatgggc aaatattata cgcaaggcga
     1681 caaggtgctg atgccgctgg cgattcaggt tcatcatgcc gtttgtgatg ggcttccatg
     1741 tcggcagaat gcttaatgaa ttacacagta ctgcgatgag tggcagggcg gggcgtaaac
     1801 gcgtggatcc ggcttactaa aagccagata acagtatgcg tatttgcgcg ctgatttttg
     1861 cggtataaga atatatactg atatgtatac ccgaagtatg tcaaaaagag gtatgctatg
     1921 aagcagcgta ttacagtgac agttgacagc gacagctatc agttgctcaa ggcatatatg
     1981 atgtcaatat ctccggtctg gtaagcacaa ccatgcagaa tgaagcccgt cgtctgcgtg
     2041 ccgaacgctg gaaagcggaa aatcaggaag ggatggctga ggtcgcccgg tttattgaaa
     2101 tgaacggctc ttttgctgac gagaacaggg gctggtgaaa tgcagtttaa ggtttacacc
     2161 tataaaactt ttgctgacga gaacaggggc tggtgaaatg cagtttaagg tttacaccta
     2221 taaaagagag agccgttatc gtctgtttgt ggatgtacag agtgatatta ttgacacgcc
     2281 cgggcgacgg atggtgatcc ccctggccag tgcacgtctg ctgtcagata aagtctcccg
     2341 tgaactttac ccggtggtgc atatcgggga tgaaagctgg cgcatgatga ccaccgatat
     2401 ggccagtgtg ccggtctccg ttatcgggga agaagtggct gatctcagcc accgcgaaaa
     2461 tgacatcaaa aacgccatta acctgatgtt ctggggaata taaatgtcag gctcccttat
     2521 acacagccag tctgcaggtc gaccatagtg actggatatg ttgtgtttta cagtattatg
     2581 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt
     2641 ctcgttcagc tttcttgtac aaagtggtga tatcgactag tgtgagcaag ggcgaggaga
     2701 ccacaatggg cgtaatcaag cccgacatga agatcaagct gaagatggag ggcaacgtga
     2761 atggccacgc cttcgtgatc gagggcgagg gcgagggcaa gccctacgac ggcaccaaca
     2821 ccatcaacct ggaggtgaag gagggagccc ccctgccctt ctcctacgac attctgacca
     2881 ccgcgttcgc ctacggcaac agggccttca ccaagtaccc cgacgacatc cccaactact
     2941 tcaagcagtc cttccccgag ggctactctt gggagcgcac catgaccttc gaggacaagg
     3001 gcatcgtgaa ggtgaagtcc gacatctcca tggaggagga ctccttcatc tacgagatac
     3061 acctcaaggg cgagaacttc ccccccaacg gccccgtgat gcagaagaag accaccggct
     3121 gggacgcctc caccgagagg atgtacgtgc gcgacggcgt gctgaagggc gacgtcaagc
     3181 acaagctgct gctggagggc ggcggccacc accgcgttga cttcaagacc atctacaggg
     3241 ccaagaaggc ggtgaagctg cccgactatc actttgtgga ccaccgcatc gagatcctga
     3301 accacgacaa ggactacaac aaggtgaccg tttacgagag cgccgtggcc cgcaactccg
     3361 aacaaaaatt gataagtgag gaagatttat aagggcccga tccggctgct aacaaagccc
     3421 gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg
     3481 cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc ggatctggcg
     3541 taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga
     3601 atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     3661 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     3721 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     3781 atttagagct ttacggcacc tcgaccgcaa aaaacttgat ttgggtgatg gttcacgtag
     3841 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     3901 tagtggactc ttgttccaaa ctggaacaac actcaaccct atcgcggtct attcttttga
     3961 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaat
     4021 atttaacgcg aattttaaca aaatattaac gtttacaatt tcgcctgatg cggtattttc
     4081 tccttacgca tctgtgcggt atttcacacc gcatacaggt ggcacttttc ggggaaatgt
     4141 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     4201 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     4261 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     4321 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     4381 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     4441 aatgatgagc acttttaaag ttctgctatg tgatacacta ttatcccgta ttgacgccgg
     4501 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     4561 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     4621 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     4681 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     4741 ggagctgaat gaagccatac caaacgacga gagtgacacc acgatgcctg tagcaatgcc
     4801 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     4861 aatagactga atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     4921 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     4981 agcactgggg ccagatggta agcgctcccg tatcgtagtt atctacacga cggggagtca
     5041 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     5101 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     5161 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     5221 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     5281 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     5341 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     5401 cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa
     5461 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     5521 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     5581 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     5641 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     5701 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     5761 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     5821 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     5881 ggccttttta cggttcctgg gcttttgctg gccttttgct cacatgttct ttcctgcgtt
     5941 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     6001 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     6061 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agg
//
