LOCUS       sequence-149378-        5566 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Single fluorescent protein biosensor for maltose.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5566)
  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 5566)
  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-Mal-B2
FEATURES             Location/Qualifiers
     source          1..5566
                     /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             2199..2450
                     /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     2387..2408
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2514..2535)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3363..3372
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     3480..3503
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             3525..3542
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3560..3577)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      3730..3816
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3908..3935
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             complement(4041..4856)
                     /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(4041..4856)
                     /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     4764..4783
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4848..4867
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     5467..5486
                     /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 gaagaaggta
     1681 aactggtaat ctggattaac ggcgataaag gctataacgg actcgctgaa gtcggtaaga
     1741 aattcgagaa agataccgga attaaagtca ccgttgagca tccggataaa ctggaagaga
     1801 aattcccaca ggttgcggca actggcgatg gccctgacat tatcttctgg gcacacgacc
     1861 gctttggtgg ctacgctcaa tctggcctgt tggctgaaat caccccggac aaagcgttcc
     1921 aggacaagct gtatccgttt acctgggatg ccgtacgtta caacggcaag ctgattgctt
     1981 acccgatcgc tgttgaagcg ttatcgctga tttataacaa agatctgctg ccgaacccgc
     2041 caaaaacctg ggaagagatc ccggcgctgg ataaagaact gaaagcgaaa ggtaagagcg
     2101 cgctgatgtt caacctgcaa gaaccgtact tcacctggcc gctgattgct gctgacgggg
     2161 gttatgcgtt cgctacttat aacgtcttta tcatggccga caagcagaag aacggcatca
     2221 aggcgaactt caagatccgc cacaacatcg aggacggcgg cgtgcagctc gcctatcact
     2281 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     2341 gcgtgcagtc caaactgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     2401 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag ggcggtaccg
     2461 gagggagcat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     2521 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     2581 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     2641 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     2701 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacatt caggagcgca
     2761 ccatcttctt caaggacgac ggcaactata agacacgcgc tgaggttaag ttcgagggcg
     2821 acactctggt taaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     2881 tgggccataa gcttgaatat aacttcaaca gtcgtaagta tgaaaacggc aagtacgaca
     2941 ttaaagacgt gggcgtggat aacgctggcg cgaaagcggg tctgaccttc ctggttgacc
     3001 tgattaaaaa caaacacatg aatgcagaca ccgattactc catcgcagaa gctgccttta
     3061 ataaaggcga aacagcgatg accatcaacg gcccgtgggc atggtccaac atcgacacca
     3121 gcaaagtgaa ttatggtgta acggtactgc cgaccttcaa gggtcaacca tccaaaccgt
     3181 tcgttggcgt gctgagcgca ggtattaacg ccgccagtcc gaacaaagag ctggcgaaag
     3241 agttcctcga aaactatctg ctgactgatg aaggtctgga agcggttaat aaagacaaac
     3301 cgctgggtgc cgtagcgctg aagtcttacg aggaagagtt ggcgaaagat ccacgtattg
     3361 ccgccaccat ggaaaacgcc cagaaaggtg aaatcatgcc gaacatcccg cagatgtccg
     3421 ctttctggta tgccgtgcgt actgcggtga tcaacgccgc cagcggtcgt cagactgtcg
     3481 atgaagccct gaaagacgcg cagactcgta tcaccaagag cggtcaccat caccatcacc
     3541 attaagagaa gattttcagc ctgatacaga ttaaatcaga acgcagaagc ggtctgataa
     3601 aacagaattt gcctggcggc agtagcgcgg tggtcccacc tgaccccatg ccgaactcag
     3661 aagtgaaacg ccgtagcgcc gatggtagtg tggggtatcc ccatgcgaga gtagggaact
     3721 gccaggcatc aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt
     3781 tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc cgggagcgga tttgaacgtt
     3841 gcgaagcaac ggcccggagg gtggcgggca ggacgcccgc cataaactgc caggcatcaa
     3901 attaagcaga aggccatcct gacggatggc cttttctgaa aggaggaact atatccggat
     3961 atccacagga cgggtgtggt cgccatgatc gcgtagtcaa ggtttcttag acgtcaggtg
     4021 gcacttttcg gggaaatgtg ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta
     4081 ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa
     4141 aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc gattccgact
     4201 cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt atcaagtgag
     4261 aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagtttatg catttctttc
     4321 cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa
     4381 ccgttattca ttcgtgattg cgcctgagcg aggcgaaata cgcgatcact gttaaaagga
     4441 caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata
     4501 ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttgcc ggggatcgca
     4561 gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt cggaagaggc
     4621 ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt ggcaacgcta
     4681 cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa tcgatagatt
     4741 gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa atcagcatcc
     4801 atgttggaat ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg gctcataaca
     4861 ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgacca aaatccctta
     4921 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcaag
     4981 ggatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     5041 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     5101 cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa
     5161 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     5221 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     5281 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     5341 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     5401 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     5461 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     5521 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaa
//
