LOCUS       sequence-111220-        6094 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 6094)
  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 6094)
  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: pFLIP32
FEATURES             Location/Qualifiers
     source          1..6094
                     /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"
     primer_bind     complement(2706..2727)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2967..2986)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3314..3335
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3351..3380
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3439..3457)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3453..3500
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3597..4052
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3684..3703)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3894..3915
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4133..4237
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4238..4306,4307..5098)
                     /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      5269..5857
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5758..5777
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6011..6028
                     /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 tctgttcacc ggggtggtgc
     2701 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     2761 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     2821 tgcccgtgcc ctggcccacc ctcgtgacca ccctgacctg gggcgtgcag tgcttcagcc
     2881 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     2941 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     3001 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     3061 acggcaacat cctggggcac aagctggagt acaactacat cagccacaac gtctatatca
     3121 ccgccgacaa gcagaagaac ggcatcaagg ccaacttcaa gatccgccac aacatcgagg
     3181 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     3241 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     3301 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc gaacaaaaat
     3361 tgataagtga ggaagattta taagggcccg atccggctgc taacaaagcc cgaaaggaag
     3421 ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     3481 gggtcttgag gggttttttg ctgaaaggag gaactatatc cggatctggc gtaatagcga
     3541 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc
     3601 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     3661 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     3721 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagagc
     3781 tttacggcac ctcgaccgca aaaaacttga tttgggtgat ggttcacgta gtgggccatc
     3841 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     3901 cttgttccaa actggaacaa cactcaaccc tatcgcggtc tattcttttg atttataagg
     3961 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa tatttaacgc
     4021 gaattttaac aaaatattaa cgtttacaat ttcgcctgat gcggtatttt ctccttacgc
     4081 atctgtgcgg tatttcacac cgcatacagg tggcactttt cggggaaatg tgcgcggaac
     4141 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     4201 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     4261 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     4321 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     4381 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     4441 cacttttaaa gttctgctat gtgatacact attatcccgt attgacgccg ggcaagagca
     4501 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     4561 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     4621 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     4681 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     4741 tgaagccata ccaaacgacg agagtgacac cacgatgcct gtagcaatgc caacaacgtt
     4801 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     4861 aatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     4921 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     4981 gccagatggt aagcgctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     5041 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     5101 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     5161 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     5221 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     5281 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     5341 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     5401 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     5461 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     5521 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     5581 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     5641 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     5701 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     5761 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     5821 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     5881 acggttcctg ggcttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     5941 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
     6001 gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc
     6061 tctccccgcg cgttggccga ttcattaatg cagg
//
