LOCUS       sequence-403702-        6677 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This is a biosensor with decreased specificity to 3OC12-HSL and 
            improved specificity to 3OHC14-HSL, containing LasR L128F-S129W 
            mutation..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6677)
  AUTHORS   Zeng M, Sarker B, Rondthaler SN, Vu V, Andrews LB
  TITLE     Identifying LasR Quorum Sensors with Improved Signal Specificity by 
            Mapping the Sequence-Function Landscape.
  JOURNAL   ACS Synth Biol. 2024 Feb 16;13(2):568-589. doi: 
            10.1021/acssynbio.3c00543. Epub 2024 Jan 11.
  PUBMED    38206199
REFERENCE   2  (bases 1 to 6677)
  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"; doi: 
            "10.1021/acssynbio.3c00543"; journalName: "ACS Synth Biol"; date: 
            "2024-02-16- 16"; volume: "13"; issue: "2"; pages: "568-589"
COMMENT     Sequence Label: pMZ103-rbs1-m8
FEATURES             Location/Qualifiers
     source          1..6677
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(399..1214)
                     /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(399..1214)
                     /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     1122..1141
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1206..1225
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     misc_RNA        1461..1510
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(1560..1562,1563..1565,1566..2279)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKLHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1605..1626)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1866..1885)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2213..2234
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        2361..2395
                     /label=BBa_J23105
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.24"
     RBS             2404..2415
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             3271..3282
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             3400..4482
                     /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     complement(3419..3438)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4501..5124
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(4574..4593)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(6058..6077)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     rep_origin      complement(join(6442..6677,1..310))
                     /direction=LEFT
                     /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     complement(6522..6541)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgcacgaac cccccgttca gtccgaccgc tgcgccttat ccggtaacta tcgtcttgag
       61 tccaacccgg aaagacatgc aaaagcacca ctggcagcag ccactggtaa ttgatttaga
      121 ggagttagtc ttgaagtcat gcgccggtta aggctaaact gaaaggacaa gttttggtga
      181 ctgcgctcct ccaagccagt tacctcggtt caaagagttg gtagctcaga gaaccttcga
      241 aaaaccgccc tgcaaggcgg ttttttcgtt ttcagagcaa gagattacgc gcagaccaaa
      301 acgatctcaa gaagatcatc ttattaaggg gtctgacgct cagtggaacg aaaaatcaat
      361 ctaaagtata tatgagtaaa cttggtctga cagttacctt agaaaaactc atcgagcatc
      421 aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt
      481 ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat
      541 cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa
      601 ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa
      661 agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa
      721 tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg
      781 cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact
      841 gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct
      901 gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc
      961 ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta
     1021 acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc
     1081 ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac
     1141 ccatataaat cagcatccat gttggaattt aatcgcggcc tggagcaaga cgtttcccgt
     1201 tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt
     1261 catgatgata tatttttatc ttgtgcaatg tacatcagag attttgagac acaaccaatt
     1321 attgaaggcc tccctaacgg ggggcctttt tttgtttctg gactcccgct tttcgagcct
     1381 agcaagggtc cgggttcacc gaaatctatc tcatttgcta gttataaaat tatgaaattt
     1441 gcgtaaattc ttcactgaag ctgtcaccgg atgtgctttc cggtctgatg agtccgtgag
     1501 gacgaaacag cctctacaaa taattttgtt taatactaga gaaagagggg aaatactaga
     1561 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     1621 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     1681 gcaagctgac cctgaagttc atctgcacca caggcaagct gcccgtgccc tggcccaccc
     1741 tcgtgaccac cttcggctac ggcctgcaat gcttcgcccg ctaccccgac cacatgaagc
     1801 tgcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     1861 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     1921 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     1981 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     2041 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     2101 accactacca gcagaacacc ccaatcggcg acggccccgt gctgctgccc gacaaccact
     2161 accttagcta ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     2221 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtaac
     2281 tcggtaccaa attccagaaa agaggcctcc cgaaaggggg gccttttttc gttttggtcc
     2341 aatggtcacc atatatcaag tttacggcta gctcagtcct aggtactatg ctagctacta
     2401 gagaaagagg agaaatacta gatggctgaa gcgcaaaatg atcccctgct gccgggatac
     2461 tcgtttaatg cccatctggt ggcgggttta acgccgattg aggccaacgg ttatctcgat
     2521 ttttttatcg accgaccgct gggaatgaaa ggttatattc tcaatctcac cattcgcggt
     2581 cagggggtgg tgaaaaatca gggacgagaa tttgtttgcc gaccgggtga tattttgctg
     2641 ttcccgccag gagagattca tcactacggt cgtcatccgg aggctcgcga atggtatcac
     2701 cagtgggttt actttcgtcc gcgcgcctac tggcatgaat ggcttaactg gccgtcaata
     2761 tttgccaata cggggttctt tcgcccggat gaagcgcacc agccgcattt cagcgacctg
     2821 tttgggcaaa tcattaacgc cgggcaaggg gaagggcgct attcggagct gctggcgata
     2881 aatctgcttg agcaattgtt actgcggcgc atggaagcga ttaacgagtc gctccatcca
     2941 ccgatggata atcgggtacg cgaggcttgt cagtacatca gcgatcacct ggcagacagc
     3001 aattttgata tcgccagcgt cgcacagcat gtttgcttgt cgccgtcgcg tctgtcacat
     3061 cttttccgcc agcagttagg gattagcgtc ttaagctggc gcgaggacca acgtatcagc
     3121 caggcgaagc tgcttttgag caccacccgg atgcctatcg ccaccgtcgg tcgcaatgtt
     3181 ggttttgacg atcaactcta tttctcgcgg gtatttaaaa aatgcaccgg ggccagcccg
     3241 agcgagttcc gtgccggtta ataaccaatt attgaaggcc gctaacgcgg cctttttttg
     3301 tttctgatct cccaatggcg gcgcgccatc gaatggcgca aaacctttcg cggtatggca
     3361 tgatagcgcc cggaagagag tcaattcagg gtggtgaata tgaaaccagt aacgttatac
     3421 gatgtcgcag agtatgccgg tgtctcttat cagaccgttt cccgcgtggt gaaccaggcc
     3481 agccacgttt ctgcgaaaac gcgggaaaaa gtggaagcgg cgatggcgga gctgaattac
     3541 attcccaacc gcgtggcaca acaactggcg ggcaaacagt cgttgctgat tggcgttgcc
     3601 acctccagtc tggccctgca cgcgccgtcg caaattgtcg cggcgattaa atctcgcgcc
     3661 gatcaactgg gtgccagcgt ggtggtgtcg atggtagaac gaagcggcgt cgaagcctgt
     3721 aaagcggcgg tgcacaatct tctcgcgcaa cgcgtcagtg ggctgatcat taactatccg
     3781 ctggatgacc aggatgccat tgctgtggaa gctgcctgca ctaatgttcc ggcgttattt
     3841 cttgatgtct ctgaccagac acccatcaac agtattattt tctcccatga ggacggtacg
     3901 cgactgggcg tggagcatct ggtcgcattg ggtcaccagc aaatcgcgct gttagcgggc
     3961 ccattaagtt ctgtctcggc gcgtctgcgt ctggctggct ggcataaata tctcactcgc
     4021 aatcaaattc agccgatagc ggaacgggaa ggcgactgga gtgccatgtc cggttttcaa
     4081 caaaccatgc aaatgctgaa tgagggcatc gttcccactg cgatgctggt tgccaacgat
     4141 cagatggcgc tgggcgcaat gcgcgccatt accgagtccg ggctgcgcgt tggtgcggat
     4201 atctcggtag tgggatacga cgataccgaa gatagctcat gttatatccc gccgttaacc
     4261 accatcaaac aggattttcg cctgctgggg caaaccagcg tggaccgctt gctgcaactc
     4321 tctcagggcc aggcggtgaa gggcaatcag ctgttgccag tctcactggt gaaaagaaaa
     4381 accaccctgg cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
     4441 cagctggcac gacaggtttc ccgactggaa agcgggcagt gacaatccag gaggaaaaaa
     4501 atgtccagat tagataaaag taaagtgatt aacagcgcat tagagctgct taatgaggtc
     4561 ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc taggtgtaga gcagcctaca
     4621 ttgtattggc atgtaaaaaa taagcgggct ttgctcgacg ccttagccat tgagatgtta
     4681 gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga ttttttacgt
     4741 aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc aaaagtacat
     4801 ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt agccttttta
     4861 tgccaacaag gtttttcact agagaatgca ttatatgcac tcagcgctgt ggggcatttt
     4921 actttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga aagggaaaca
     4981 cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt tgatcaccaa
     5041 ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt agaaaaacaa
     5101 cttaaatgtg aaagtgggtc ctaatctaat ttaacaaaag gttttccaat ttatggcctt
     5161 ggttgacggt tttcttgagc tggaacgctc aagtggaaaa ttggagtgga gcgccatcct
     5221 gcagaagatg gcgagcgacc ttggattctc gaagatcctg ttcggcctgt tgcctaagga
     5281 cagccaggac tacgagaacg ccttcatcgt cggcaactac ccggccgcct ggcgcgagca
     5341 ttacgaccgg gctggctacg cgcgggtcga cccgacggtc agtcactgta cccagagcgt
     5401 actgccgatt ttctgggaac cgtccatcta ccagacgcga aagcagcacg agttcttcga
     5461 ggaagcctcg gccgccggcc tggtgtatgg gctgaccatg ccgctgcatg gtgctcgcgg
     5521 cgaactcggc gcgttttggc tcagcgtgga agcggaaaac cgggccgagg ccaaccgttt
     5581 catggagtcg gtcctgccga ccctgtggat gctcaaggac tacgcactgc agagcggtgc
     5641 cggactggcc ttcgaacatc cggtcagcaa accggtggtt ctgaccagcc gggagaagga
     5701 agtgttgcag tggtgcgcca tcggcaagac cagttgggag atatcggtta tctgcaactg
     5761 ctcggaagcc aatgtgaact tccatatggg aaatattcgg cggaagttcg gtgtgacctc
     5821 ccgccgcgta gcggccatta tggccgttaa tttgggtctt attactctct gaccataatt
     5881 ggtaacgaat cagacaattg acggctcgag ggagtagcat agggtttgca gaatccctgc
     5941 ttcgtccatt tgacaggcac attatgcatc gatgataagc tgtcaaacat gagcagatcc
     6001 tctacgccgg acgcatcgtg gccggcatca ccggcgccac aggtgcggtt gctggcgcct
     6061 atatcgccga catcaccgat ggggaagatc gggctcgcca cttcgggctc atgagcaaat
     6121 attttatctg aggtgcttcc tcgctcactg actcgctgca cgaggcagac ctcagcgcta
     6181 gcggagtgta tactggctta ctatgttggc actgatgagg gtgtcagtga agtgcttcat
     6241 gtggcaggag aaaaaaggct gcaccggtgc gtcagcagaa tatgtgatac aggatatatt
     6301 ccgcttcctc gctcactgac tcgctacgct cggtcgttcg actgcggcga gcggaaatgg
     6361 cttacgaacg gggcggagat ttcccggaag atgccaggaa gatacttaac agggaagtga
     6421 gagggccgcg gcaaagccgt ttttccatag gctccgcccc cctgacaagc atcacgaaat
     6481 ctgacgctca aatcagtggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     6541 ccctggcggc tccctcgtgc gctctcctgt tcctgccttt cggtttaccg gtgtcattcc
     6601 gctgttatgg ccgcgtttgt ctcattccac gcctgacact cagttccggg taggcagttc
     6661 gctccaagct ggactgt
//
