LOCUS       sequence-244017-        6539 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mercury sensor with an optimized HrpRS-based amplifier. pSB3K3 
            carrying J109-32merR-t-PmerT-30hrpR-30hrpS-t-PhrpLE-30gfp-t.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6539)
  AUTHORS   Wan X, Volpetti F, Petrova E, French C, Maerkl SJ, Wang B
  TITLE     Cascaded amplifying circuits enable ultrasensitive cellular sensors 
            for toxic metals.
  JOURNAL   Nat Chem Biol. 2019 May;15(5):540-548. doi: 
            10.1038/s41589-019-0244-3. Epub 2019 Mar 25.
  PUBMED    30911179
REFERENCE   2  (bases 1 to 6539)
  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: 
            "10.1038/s41589-019-0244-3"; journalName: "Nat Chem Biol"; date: 
            "2019-05"; volume: "15"; issue: "5"; pages: "540-548"
COMMENT     Sequence Label: pXWJ109HgAmp30E
FEATURES             Location/Qualifiers
     source          1..6539
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..22
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     promoter        23..57
                     /label=BBa_J23109
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.04"
     terminator      536..607
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      623..650
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             733..744
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1655..1666
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     RBS             1729..1740
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      2667..2738
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2754..2781
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             3015..3026
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             3033..3746
                     /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="MRKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(3063..3091)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     3686..3706
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      3769..3840
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3856..3883
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    3890..3910
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     CDS             complement(4050..4865)
                     /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(4050..4865)
                     /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     4773..4792
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4857..4876
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      complement(5460..6004)
                     /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(5539..5558)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6472..6490
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(6494..6537)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
ORIGIN
        1 gaattcgcgg ccgcttctag agtttacagc tagctcagtc ctagggactg tgctagctac
       61 tagagtcaca caggaaagta ctagatggaa aataatttgg aaaacctgac cattggcgtt
      121 tttgccaagg cggccggggt caacgtggag acaatccgct tctatcagcg caagggcctg
      181 ttgcgggaac cggacaagcc ttacggcagc atccgccgct atggggaggc ggacgtggtt
      241 cgggtgaaat tcgtgaaatc ggcacagcgg ctggggttca gtctggacga gattgccgag
      301 ctgttgcggc tcgacgatgg cacccactgc gaggaggcca gcagcctggc cgaacacaag
      361 ctcaaggacg tgcgcgagaa gatggccgac ttggcgcgca tggaaaccgt gctgtctgaa
      421 ctcgtgtgcg cctgccatgc acgaaagggg aatgtttcct gcccgttgat cgcgtcacta
      481 cagggcgaag caggcctggc aaggtcagct atgccttagt actagagcca ggcatcaaat
      541 aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa
      601 cgctctctac tagagtcaca ctggctcacc ttcgggtggg cctttctgcg tttatatgga
      661 tccgttccat atcgcttgac tacgtacatg agtacggaag taaggttacg ctatccaatc
      721 ctactagaga ttaaagagga gaaatactag atgagtacag gcatcgataa ggacgtccga
      781 gagtgttggg gcgtaactgc attatcagcg ggtcatcaaa ttgcaatgaa tagcgcgttt
      841 ctggatatgg acttgctgtt gtgcggggaa accggcaccg gcaaggacac actggccaac
      901 cgcattcacg agttgtccag caggtcggga ccctttgtgg gcatgaactg cgccgccatt
      961 cccgagtcgc tggcagagag ccagttattc ggtgtggtca acggtgcatt caccggcgta
     1021 tgccgggctc gcgagggcta catagaggcc tccagtggtg gcaccttgta cctggatgaa
     1081 atcgacagca tgccgttgag cctgcaagcc aaactgctgc gtgtgttgga gagtcgaggt
     1141 atcgagcgtc tgggctcgac cgaatttatc ccggtggatc tgcggatcat tgcctcggcc
     1201 cagcggccac tggatgaact ggtggaacaa ggacttttcc gtcgcgacct gttttttcgg
     1261 ctcaacgtgc tgacgcttca cttgccagcc ttgcgcaaac gtcgtgaaca gatcctgcca
     1321 ttgttcgacc agttcaccca gggtatcgct gccgagttcg gacgtcccgc tcctgcgctg
     1381 gacagcgggc gtgtgcagct gctgctcagc cacgactggc cgggcaacat ccgcgaattg
     1441 aagtctgcgg ccaagcgctt cgtactcggc ttccccttgc tgggcgccga ccctgtggaa
     1501 gcgcttgacc ctgccacggg gctgcgcacg caaatgcgca tcatcgagaa aatgctcatc
     1561 caggatgcct tgaagcggca caggcacaat ttcgacgcgg tgcttcagga gttggagttg
     1621 ccaagacgca ccctgtatca ccgcatgaag gaactgggag ttgcagcgcc gatcgctgcg
     1681 acggccgggg tctaataaaa cgcaggtgtg gttatgctac tagagattaa agaggagaaa
     1741 tactagatga gtcttgatga aaggtttgag gatgatctgg acgaggagcg ggttccgaat
     1801 ctggggatag ttgccgaaag tatttcgcaa ctgggtatcg acgtgctgct atcgggtgag
     1861 accggcacgg gcaaagacac gattgcccga cggattcatg agatgtcagg ccgcaaaggg
     1921 cgcctggtgg cgatgaattg cgcggccatt ccggagtccc tcgccgagag cgagttattc
     1981 ggcgtggtca gcggtgccta caccggcgct gatcgctcca gagtcggtta tgtcgaagcg
     2041 gcgcagggcg gcacgctgta cctggatgag atcgatagca tgccgctgag cctgcaagcc
     2101 aaattgctga gggtgctgga aacccgagcg cttgaacggc tgggttcgac gtcgacgatc
     2161 aagctggata tctgcgtgat cgcctccgcc caatgctcgc tggacgacgc cgtcgagcgg
     2221 gggcagtttc gtcgcgatct gtattttcgc ctgaacgtcc tgacactcaa gcttcctccg
     2281 ctacgtaacc agtctgatcg catagttccc ctgttcacac gttttacggc cgccgccgcg
     2341 agggagctcg gtgttcccgt tcccgatgtt tgcccactgc tgcacaaagt gctgctgggc
     2401 cacgactggc ccggcaatat ccgtgagctc aaggcggcag ccaaacgcca tgtgctgggt
     2461 ttccccttgc tgggcgccga gccgcagggc gaagagcact tggcctgtgg gctcaaatcg
     2521 caattgcgag tgatcgaaaa agccctgatt caggagtcgc tcaagcgcca cgacaattgt
     2581 gtggattcgg taagcctgga actggacgtg ccacgccgta cgctctatcg acgcatcaaa
     2641 gaattgcaga tctaataacc aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg
     2701 cctttcgttt tatctgttgt ttgtcggtga acgctctcta ctagagtcac actggctcac
     2761 cttcgggtgg gcctttctgc gtttatatac tagaggccgg attatgtccg ctgagtgggt
     2821 cacggtcccg gatcagttcc cttgcgaagc tgaccgatgt ttttgtgcca aaagctgttg
     2881 tggcaaaaaa cggtttgcgc aaagttttgt attacaaaga atttcacatt ttaaaatatc
     2941 tttataaatc aatcagttat ttctattttc aagctggcac ggttattgct atagggcttg
     3001 tactactaga gattaaagag gagaaatact agatgcgtaa aggagaagaa cttttcactg
     3061 gagttgtccc aattcttgtt gaattagatg gtgatgttaa tgggcacaaa ttttctgtca
     3121 gtggagaggg tgaaggtgat gcaacatacg gaaaacttac ccttaaattt atttgcacta
     3181 ctggaaaact acctgttcca tggccaacac ttgtcactac tttcggttat ggtgttcaat
     3241 gctttgcgag atacccagat catatgaaac agcatgactt tttcaagagt gccatgcccg
     3301 aaggttatgt acaggaaaga actatatttt tcaaagatga cgggaactac aagacacgtg
     3361 ctgaagtcaa gtttgaaggt gatacccttg ttaatagaat cgagttaaaa ggtattgatt
     3421 ttaaagaaga tggaaacatt cttggacaca aattggaata caactataac tcacacaatg
     3481 tatacatcat ggcagacaaa caaaagaatg gaatcaaagt taacttcaaa attagacaca
     3541 acattgaaga tggaagcgtt caactagcag accattatca acaaaatact ccaattggcg
     3601 atggccctgt ccttttacca gacaaccatt acctgtccac acaatctgcc ctttcgaaag
     3661 atcccaacga aaagagagac cacatggtcc ttcttgagtt tgtaacagct gctgggatta
     3721 cacatggcat ggatgaacta tacaaataat aatactagag ccaggcatca aataaaacga
     3781 aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc
     3841 tactagagtc acactggctc accttcgggt gggcctttct gcgtttatat actagtagcg
     3901 gccgctgcag gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     3961 ggtatcagct cactcaaagg cggtaatctc gagtcccgtc aagtcagcgt aatgctctgc
     4021 cagtgttaca accaattaac caattctgat tagaaaaact catcgagcat caaatgaaac
     4081 tgcaatttat tcatatcagg attatcgata ccatattttt gaaaaagccg tttctgtaat
     4141 gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg
     4201 attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta
     4261 tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa aagcttatgc
     4321 atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca
     4381 tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg
     4441 ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca
     4501 tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg
     4561 gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc
     4621 ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg
     4681 gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat
     4741 cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa
     4801 tcagcatcca tgttggaatt taatcgcggc ctggagcaag acgtttcccg ttgaatatgg
     4861 ctcataacac cccttgtatt actgtttatg taagcagaca gttttattgt tcatgatgat
     4921 atatttttat cttgtgcaat gtaacatcag agattttgag acacaacgtg gctttgttga
     4981 ataaatcgaa cttttgctga gttgaaggat cagatcacgc atcttcccga caacgcagac
     5041 cgttccgtgg caaagcaaaa gttcaaaatc accaactggt ccacctacaa caaagctctc
     5101 atcaaccgtg gctccctcac tttctggctg gatgatgggg cgattcaggc ctggtatgag
     5161 tcagcaacac cttcttcacg aggcagacct cagcgctagc ggagtgtata ctggcttact
     5221 atgttggcac tgatgagggt gtcagtgaag tgcttcatgt ggcaggagaa aaaaggctgc
     5281 accggtgcgt cagcagaata tgtgatacag gatatattcc gcttcctcgc tcactgactc
     5341 gctacgctcg gtcgttcgac tgcggcgagc ggaaatggct tacgaacggg gcggagattt
     5401 cctggaagat gccaggaaga tacttaacag ggaagtgaga gggccgcggc aaagccgttt
     5461 ttccataggc tccgccccct gacaagcatc acgaaatctg acgctcaaat cagtggtggc
     5521 gaaacccgac aggactataa agataccagg cgtttccccc tggcggctcc ctcgtgcgct
     5581 ctcctgttcc tgcctttcgg tttaccggtg tcattccgct gttatggccg cgtttgtctc
     5641 attccacgcc tgacactcag ttccgggtag gcagttcgct ccaagctgga ctgtatgcac
     5701 gaaccccccg ttcagtccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5761 ccggaaagac atgcaaaagc accactggca gcagccactg gtaattgatt tagaggagtt
     5821 agtcttgaag tcatgcgccg gttaaggcta aactgaaagg acaagttttg gtgactgcgc
     5881 tcctccaagc cagttacctc ggttcaaaga gttggtagct cagagaacct tcaaaaaacc
     5941 gccctgcaag gcggtttttt cgttttcaga gcaagagatt acgcgcagac caaaacgatc
     6001 tcaagaagat catcttatta aggggtctga cgctcagtgg aacgaaaact cacgttaagg
     6061 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     6121 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     6181 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     6241 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     6301 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     6361 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattccatg
     6421 gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt
     6481 atcacgaggc agaatttcag ataaaaaaaa tccttagctt tcgctaagga tgatttctg
//
