LOCUS       sequence-149105-        5695 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Single fluorescent protein biosensor for trehalose.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5695)
  AUTHORS   Nadler DC, Morgan SA, Flamholz A, Kortright KE, Savage DF
  TITLE     Rapid construction of metabolite biosensors using domain-insertion 
            profiling.
  JOURNAL   Nat Commun. 2016 Jul 29;7:12266. doi: 10.1038/ncomms12266.
  PUBMED    27470466
REFERENCE   2  (bases 1 to 5695)
  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: "Nat 
            Commun."; date: "2016-07-29"; volume: "7:12266. doi"; pages: " 
            10.1038/ncomms12266"
COMMENT     Sequence Label: pTKEI-Tre-C04
FEATURES             Location/Qualifiers
     source          1..5695
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     154..171
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(379..1461)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     1423..1442
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1462..1539)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(1584..1589,1590..1606,1607..1613)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1621..1637
                     /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)."
     CDS             2697..2948
                     /codon_start=1
                     /product="C-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VC155
                     /translation="DKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGPVLLPDNH
                     YLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYK"
     primer_bind     2885..2906
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3012..3033)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             3654..3671
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3689..3706)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      3859..3945
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4037..4064
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             complement(4170..4985)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(4170..4985)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4893..4912
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4977..4996
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     5596..5615
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
       61 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
      121 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      181 gagcgccacg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc tagccgggtc
      241 ctcaacgaca ggagcacgat catgctagtc atgccccgcg cccaccggaa ggagctgact
      301 gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt
      361 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
      421 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt
      481 ttttcttttc accagtgaca cgggcaacag ctgattgccc ttcaccgcct ggccctgaga
      541 gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt
      601 ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat
      661 atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg
      721 atcgttggca accagcatcg cagtgggaac gataccctca ttcagcattt gcatggtttg
      781 ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt
      841 gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg
      901 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
      961 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
     1021 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
     1081 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
     1141 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
     1201 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
     1261 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
     1321 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
     1381 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
     1441 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
     1501 tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct
     1561 tatgcgactc cgatcccgca aatttgacaa ttaatcatcc ggctcgtata atgtgtggaa
     1621 ttgtgagcgg ataacaattt cacacaagga gatataccat gtccaaaatc gaggaaggta
     1681 aaattgtgtt tgcagtgggt ggcgccccga atgaaatcga atattggaaa ggtgttattg
     1741 ccgaattcga gaaaaaatat cctggggtta cagtggagct gaaacgccag gccaccgata
     1801 ccgaacagcg tcgtctggat ctggttaatg cgcttcgcgg gaaaagctcc gaccctgatg
     1861 tattcttgat ggacgtcgcg tggctgggtc agttcatcgc ttctggttgg ctggaacccc
     1921 tggacgatta tgttcaaaaa gacaattatg acctgagcgt gttctttcag tcggtgatta
     1981 accttgcgga caaacagggc ggcaaactgt atgccctgcc ggtgtatatt gatgcaggtc
     2041 tgctgtacta tcgtaaagat cttcttgaaa aatacggcta ttctaagccg ccggaaacat
     2101 ggcaggaact ggtcgagatg gcccaaaaaa ttcagtcggg ggagcgcgaa accaacccaa
     2161 atttttgggg ttttgtttgg cagggcaaac aatacgaggg cctggtgtgc gacttcgtgg
     2221 agtacgtgta ctcaaacggc ggaagcttag gtgaatttaa agacggcaag tgggtcccga
     2281 ctttgaacaa accggaaaat gttgaggccc tgcaattcat ggtggatctg atccacaagt
     2341 ataagatttc gcctccaaac acctacacgg aaatgacgga agaaccggta cgcctgatgt
     2401 ttcaacaggg caacgcggcg ttcgagcgta attggccata tgcttggggc ctgcataatg
     2461 ctgacgattc tccagtgaaa ggcaaagtcg gtgtggcccc gttgccacat tttccgggcc
     2521 ataaaagcgc ggctaccctg ggtggttggc acattggtat tagcaagtat agcgacaata
     2581 aagccttggc gtgggaattt gtgaaattcg tagaaagtta tagcgtccag aaaggatttg
     2641 cgatgaattt gggttggaat ccgggcgacc gctcttataa cgtctttatc atggccgaca
     2701 agcagaagaa cggcatcaag gcgaacttca agatccgcca caacatcgag gacggcggcg
     2761 tgcagctcgc ctatcactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     2821 ccgacaacca ctacctgagc gtgcagtcca aactgagcaa agaccccaac gagaagcgcg
     2881 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     2941 tgtacaaggg cggtaccgga gggagcatgg tgagcaaggg cgaggagctg ttcaccgggg
     3001 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg
     3061 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     3121 gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct
     3181 tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     3241 gctacattca ggagcgcacc atcttcttca aggacgacgg caactataag acacgcgctg
     3301 aggttaagtt cgagggcgac actctggtta accgcatcga gctgaagggc atcgacttca
     3361 aggaggacgg caacatcctg ggccataagc ttgaatataa cttcaaccgc agccgcggtc
     3421 gcgtagacgt atacgacgat ccggcggtgg tgtccaaaag cccccatctg aaagagctgc
     3481 gtgcggtgtt tgaaaacgcg gtcccacgcc caatcgtacc gtattacccg caactgagtg
     3541 aaatcatcca gaaatacgtt aactccgccc ttgcaggcaa aatctccccg caggaagcac
     3601 tggataaagc tcaaaaagaa gccgaagaac tggtaaagca gtattcaagc ggtcaccatc
     3661 accatcacca ttaagagaag attttcagcc tgatacagat taaatcagaa cgcagaagcg
     3721 gtctgataaa acagaatttg cctggcggca gtagcgcggt ggtcccacct gaccccatgc
     3781 cgaactcaga agtgaaacgc cgtagcgccg atggtagtgt ggggtatccc catgcgagag
     3841 tagggaactg ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt
     3901 tttatctgtt gtttgtcggt gaacgctctc ctgagtagga caaatccgcc gggagcggat
     3961 ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag gacgcccgcc ataaactgcc
     4021 aggcatcaaa ttaagcagaa ggccatcctg acggatggcc ttttctgaaa ggaggaacta
     4081 tatccggata tccacaggac gggtgtggtc gccatgatcg cgtagtcaag gtttcttaga
     4141 cgtcaggtgg cacttttcgg ggaaatgtgt tagaaaaact catcgagcat caaatgaaac
     4201 tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat
     4261 gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg
     4321 attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta
     4381 tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc
     4441 atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca
     4501 tcaaccaaac cgttattcat tcgtgattgc gcctgagcga ggcgaaatac gcgatcactg
     4561 ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca
     4621 tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc tgttttgccg
     4681 gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc
     4741 ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg
     4801 gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat
     4861 cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa
     4921 tcagcatcca tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg
     4981 ctcataacac cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa
     5041 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     5101 atcttcaagg gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     5161 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     5221 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
     5281 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     5341 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     5401 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     5461 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5521 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5581 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5641 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaa
//
