LOCUS       sequence-403606-        8186 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A C4-HSL sensor for B. subtilis on a shuttle vector for B. subtilis 
            and E. coli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8186)
  AUTHORS   Zeng M, Sarker B, Howitz N, Shah I, Andrews LB
  TITLE     Synthetic Homoserine Lactone Sensors for Gram-Positive Bacillus 
            subtilis Using LuxR-Type Regulators.
  JOURNAL   ACS Synth Biol. 2023 Dec 11. doi: 10.1021/acssynbio.3c00504.
  PUBMED    38079538
REFERENCE   2  (bases 1 to 8186)
  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: "ACS Synth 
            Biol. 2023 Dec 11. doi: 10.1021/acssynbio.3c00504."
COMMENT     Sequence Label: pMZ3012
FEATURES             Location/Qualifiers
     source          1..8186
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(2118..2137)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2129..2944
                     /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             2129..2944
                     /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     complement(2202..2221)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3033..3578
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     3479..3498
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3695..3712
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(4572..5654)
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5616..5635
                     /label=LacI-R
                     /note="LacI, reverse primer"
     terminator      complement(5902..5936)
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     CDS             complement(7051..7764)
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFAYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(7091..7111)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     7706..7734
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     misc_RNA        complement(7821..7870)
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     terminator      complement(7961..7988)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(8007..8093)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 aaaaaaacac aaaagaccac attttttaat gtggtctttt attcttcaac taaagcaccc
       61 attagttcaa caaacgaaaa ttggataaag tgggatattt ttaaaatata tatttatgtt
      121 acagtaatat tgacttttaa aaaaggattg attctaatga agaaagcaga caagtaagcc
      181 tcctaaattc actttagata aaaatttagg aggcatatca aatgaacttt aataaaattg
      241 atttagacaa ttggaagaga aaagagatat ttaatcatta tttgaaccaa caaacgactt
      301 ttagtataac cacagaaatt gatattagtg ttttataccg aaacataaaa caagaaggat
      361 ataaatttta ccctgcattt attttcttag tgacaagggt gataaactca aatacagctt
      421 ttagaactgg ttacaatagc gacggagagt taggttattg ggataagtta gagccacttt
      481 atacaatttt tgatggtgta tctaaaacat tctctggtat ttggactcct gtaaagaatg
      541 acttcaaaga gttttatgat ttataccttt ctgatgtaga gaaatataat ggttcgggga
      601 aattgtttcc caaaacacct atacctgaaa atgctttttc tctttctatt attccatgga
      661 cttcatttac tgggtttaac ttaaatatca ataataatag taattacctt ctacccatta
      721 ttacagcagg aaaattcatt aataaaggta attcaatata tttaccgcta tctttacagg
      781 tacatcattc tgtttgtgat ggttatcatg caggattgtt tatgaactct attcaggaat
      841 tgtcagatag gcctaatgac tggcttttat aatatgagat aatgccgact gtacttttta
      901 cagtcggttt tctaatgtca ctaacctgcc ccgccagaaa acaataaacc cttgcatagg
      961 gggaatccgt ttaggctggg cggtgatagc ttctcgttca ggcagtacgc ctcttttctt
     1021 ttccagacct gagggaggcg gaaatggtgt gaggttcccg gggaaaagcc aaataggcga
     1081 tcgcgggagt gctttatttg aagatcaggc tatcactgcg gtcaatagat ttcacaatgt
     1141 gatggctgga cagcctgagg aactctcgaa cccgaatgga aacaaccaga tatttatgaa
     1201 tcagcgcggc tcacatggcg ttgtgctggc aaatgcaggt tcatcctctg tctctatcaa
     1261 tacggcaaca aaattgcctg atggcaggta tgacaataaa gctggagcgg gttcatttca
     1321 agtgaacgat ggtaaactga caggcacgat caatgccagg tctgtagctg tgctttatcc
     1381 tgatgatatt gcaaaagcgc ctcatgtttt ccttgagaat tacaaaacag gtgtaacaca
     1441 ttctttcaat gatcaactga cgattacctt gcgtgcagat gcgaatacaa caaaagccgt
     1501 ttatcaaatc aataatggac cagagacggc gtttaaggat ggagatcaat tcacaatcgg
     1561 aaaaggagat ccatttggca aaacatacac catcatgtta aaaggaacga acagtgatgg
     1621 tgtaacgagg accgagaaat acagttttgt taaaagagat ccagcgtcgg ccaaaaccat
     1681 cggctatcaa aatccgaatc attggagcca ggtaaatgct tatatctata aacatgatgg
     1741 gagccgagta attgaattga ccggatcttg gcctggaaaa ccaatgacta aaaatgcaga
     1801 cggaatttac acgctgacgc tgcctgcgga cacggataca accaacgcaa aagtgatttt
     1861 taataatggc agcgcccaag tgcccggtca gaatcagcct ggctttgatt acgtgctaaa
     1921 tggtttatat aatgactcgg gcttaagcgg ttctcttccc cattgaccca gaagcgggag
     1981 accagaaaca aaaaaaggcc ccccgttagg gaggccttca ataattggtt gtgtctcaaa
     2041 atctctgatg tacattgcac aagataaaaa tatatcatca tgaacaataa aactgtctgc
     2101 ttacataaac agtaatacaa ggggtgttat gagccatatt caacgggaaa cgtcttgctc
     2161 caggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat gggctcgcga
     2221 taatgtcggg caatcaggtg cgacaatcta tcgattgtat gggaagcccg atgcgccaga
     2281 gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg agatggtcag
     2341 actaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta tccgtactcc
     2401 tgatgatgca tggttactca ccactgcgat ccccgggaaa acagcattcc aggtattaga
     2461 agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt
     2521 gcattcgatt cctgtttgta attgtccttt taacagcgat cgcgtatttc gtctcgctca
     2581 ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg acgagcgtaa
     2641 tggctggcct gttgaacaag tctggaaaga aatgcataag cttttgccat tctcaccgga
     2701 ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg aggggaaatt
     2761 aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg atcttgccat
     2821 cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt ttcaaaaata
     2881 tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg atgagttttt
     2941 ctaaggtaac tgtcagacca agtttactca tatatacttt agattgattt ttcgttccac
     3001 tgagcgtcag accccttaat aagatgatct tcttgagatc gttttggtct gcgcgtaatc
     3061 tcttgctctg aaaacgaaaa aaccgccttg cagggcggtt tttcgaaggt tctctgagct
     3121 accaactctt tgaaccgagg taactggctt ggaggagcgc agtcaccaaa acttgtcctt
     3181 tcagtttagc cttaaccggc gcatgacttc aagactaact cctctaaatc aattaccagt
     3241 ggctgctgcc agtggtgctt ttgcatgtct ttccgggttg gactcaagac gatagttacc
     3301 ggataaggcg cagcggtcgg actgaacggg gggttcgtgc atacagtcca gcttggagcg
     3361 aactgcctac ccggaactga gtgtcaggcg tggaatgaga caaacgcggc cataacagcg
     3421 gaatgacacc ggtaaaccga aaggcaggaa caggagagcg cacgagggag ccgccagggg
     3481 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca ccactgattt gagcgtcaga
     3541 tttcgtgatg cttgtcaggg gggcggagcc tatggaaaaa cggctttgcc gcggccctct
     3601 cacttccctg ttaagtatct tcctggcatc ttccgggaaa tctccgcccc gttcgtaagc
     3661 catttccgct cgccgcagtc gaacgaccga gcgtagcgag tcagtgagcg aggaagcgga
     3721 atatatcctg tatcacatat tctgctgacg caccggtgca gccttttttc tcctgccaca
     3781 tgaagcactt cactgacacc ctcatcagtg ccaacatagt aagccagtat acactccgct
     3841 agcgctgagg tctgcctcgt gcttcaaaaa atcaaataag gagtgtcaag aatgtttgca
     3901 aaacgattca aaacctcttt actgccgtta ttcgctggat ttttattgct gtttcatttg
     3961 gttctggcag gaccggcggc tgcgagtgct gaaacggcga acaaatcgaa tgagcttaca
     4021 gcaccgtcga tcaaaagcgg aaccattctt catgcatgga attggtcgtt caatacgtta
     4081 aaacacaata tgaaggatat tcatgatgca ggatatacag ccattcagac atctccgatt
     4141 aaccaagtaa aggaagggaa tcaaggagat aaaagcatgt cgaactggta ctggctgtat
     4201 cagccgacat cgtatcaaat tggcaaccgt tacttaggta ctgaacaaga atttaaagaa
     4261 atgtgtgcag ccgctgaaga atatggcata aaggtcattg ttgacgcggt catcaatcat
     4321 accaccagtg attatgccgc gatttccaat gaggttaaga gtattccaaa ctggacacat
     4381 ggaaacacac aaattaaaaa ctggtctgat cgatgggatg tcacgcagaa ttaaatagta
     4441 cataatggat ttccttacgc gaaatacggg cagacatggc ctgcccggtt attattattt
     4501 ttgacaccag accaactggt aatggtagcg accggcgctc aggatcctaa ctcacattaa
     4561 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     4621 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt
     4681 ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc
     4741 agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttgac
     4801 ggcgggatat aacatgagct atcttcggta tcgtcgtatc ccactaccga gatatccgca
     4861 ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg
     4921 gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa
     4981 ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg
     5041 agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct
     5101 aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg
     5161 tcctcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac
     5221 gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag
     5281 ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct
     5341 tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga
     5401 gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg
     5461 ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc
     5521 agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg
     5581 ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac
     5641 gttactggtt tcatcaaaat cgtctccctc cgtttgaata tttgattgat cgtaaccaga
     5701 tgaagcactc tttccactat ccctacagtg ttatggcttg aacaatcacg aaacaataat
     5761 tggtacgtac gatctttcag ccgactcaaa catcaaatct tacaaatgta gtctttgaaa
     5821 gtattacata tgtaagattt aaatgcaacc gttttttcgg aaggaaatga tgacctcgag
     5881 caacgttctt gccattgctg cataaaaaac gcccggcggc aaccgagcgt tctgaattaa
     5941 ttaatcatcg ggaagatctt catcaccgaa acgcggcagg cagctctaga gttaacatta
     6001 cctggaccaa aacgaaaaaa gacgctctaa aagcgtctct tttctggaat ttggtaccga
     6061 gcattttaga tgaggcccag cgccgcggcg taggcggcag ccagcgtctt gttcggcgcg
     6121 tcgaacttct tctggatgtt cttgtggtgg aagttcaccg tgctctcgga gatgctcagg
     6181 atgatggcga tttccccgga actcttgccg tcggcggtcc attgcaggat ctcgcgttcg
     6241 cgatggctca ggcagaccgg gttggacatc agcatcggat gctccaggtc ggtcagcttc
     6301 tgggtcagca actcgatcat gcaacgcagc cgcaggcgga tttcctcgcg ctcgaagctg
     6361 gagatgttct gctggtcgcg cgccacggaa agcacgctga gcaaattgtt cggcgcgcgg
     6421 atcggcaagg tcgcgccgac acagaggccc caatcgcgag cctcgttcca gagcatccgg
     6481 ctctggtcga acaggctgtc gctccagacc accatttccg aggagcgcag gccgttgagg
     6541 atcgccggat ccacggcccc gtagttctgc atctggtatc gctccagcca ggccttggga
     6601 taggtgccat ggacctcggt cttgggccgg gtgaagggaa tcgtgtggcg cacgccatag
     6661 gcgtagtaat cgaagcccag gcgccgcact tccttttcca ggacggcgaa cacgccctgg
     6721 ctgtcgtgga tcggctgcat ctcgctacgc aaaccgtccc accacagcaa aaagcctccg
     6781 tcattcctca taatatttta cctcctttac ttactcccta acgtattaaa caaaattatt
     6841 tgtagaggtc ataagtttcg tcgcatttca gcgactcatc agtgggctta gcccagactt
     6901 atgacttcag tcacaactat gttcaatagt accatagaaa atcattccaa tacaaacaat
     6961 aaccatctcc agcgacctca ttggaccaaa acgaaaaaag gccccccttt cgggaggcct
     7021 cttttctgga atttggtacc gagttactta tttgtatagt tcatccatgc catgtgtaat
     7081 cccagcagct gttacaaact caagaaggac catgtgatct ctcttttcgt tgggatcttt
     7141 cgaaagggca gattggtagg acaggtaatg gttgtctggt aaaaggacag ggccatcgcc
     7201 aattggagta ttttgttgat aatggtctgc tagttgaacg cttccatctt caatgttgtg
     7261 tctaattttg aagttaactt tgattccatt cttttgtttg tctgccatga tgtatacatt
     7321 gtgtgagtta tagttgtatt ccaatttgtg tccaagaatg tttccatctt ctttaaaatc
     7381 aatacctttt aactcgattc tattaacaag ggtatcacct tcaaacttga cttcagcacg
     7441 tgtcttgtag ttcccgtcat ctttgaaaaa tatagttctt tcctgtacat aaccttcggg
     7501 catggcactc ttgaaaaagt catgctgttt catatgatct gggtatctcg caaagcattg
     7561 aagtccatac gcgaaagtag tgacaagtgt tggccatgga acaggtagtt ttccagtagt
     7621 gcaaataaat ttaagggtaa gttttccgta tgttgcatca ccttcaccct ctccactgac
     7681 agaaaatttg tgcccattaa catcaccatc taattcaaca agaattggga caactccagt
     7741 gaaaagttct tctcctttac tcatagtagt tcctccttat gtaagcttcc tctagtatta
     7801 aacaaaatta tttgtagagg ctgtttcgtc ctcacggact catcagaccg gaaagcacat
     7861 ccggtgacag cttcagacta catttattat agccaattcg aaagctaacg gtaactgcca
     7921 gatttcacag gactccaagc caagagtttg tagaaacgca aaaaggccat ccgtcaggat
     7981 ggccttctgc ttagctagag cggcggattt gtcctactca ggagagcgtt caccgacaaa
     8041 caacagataa aacgaaaggc ccagtctttc gactgagcct ttcgttttat ttgatgcctc
     8101 aagctagaga gtcgaattcc cggcaatagt tacccttatt atcaagataa gaaagaaaag
     8161 gatttttcgc tacgctcaaa tccttt
//
