LOCUS       sequence-240606-        6551 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Arsenic sensor with an optimized HrpRS-based amplifier. pSB3K3 
            carrying J109-30arsR-t-ParsR-30hrpR-30hrpS-t-PhrpLE-30gfp-t.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6551)
  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 6551)
  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: pXWJ109AsAmp30E
FEATURES             Location/Qualifiers
     source          1..6551
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        22..56
                     /label=BBa_J23109
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.04"
     RBS             68..79
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      502..573
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      589..616
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             744..755
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1666..1677
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     RBS             1740..1751
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      2678..2749
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2765..2792
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             3026..3037
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             3044..3757
                     /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(3074..3102)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     3697..3717
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      3780..3851
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3867..3894
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    3901..3921
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     CDS             complement(4061..4876)
                     /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(4061..4876)
                     /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     4784..4803
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4868..4887
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      complement(5471..6015)
                     /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(5550..5569)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6483..6501
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(6505..6548)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    join(6551,1..21)
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
ORIGIN
        1 aattcgcggc cgcttctaga gtttacagct agctcagtcc tagggactgt gctagctact
       61 agagattaaa gaggagaaat actagatgtc atttctgtta cccatccaat tgttcaaaat
      121 tcttgctgat gaaacccgtc tgggcatcgt tttactgctc agcgaactgg gagagttatg
      181 cgtctgcgat ctctgcactg ctctcgacca gtcgcagccc aagatctccc gccacctggc
      241 attgctgcgt gaaagcgggc tattgctgga ccgcaagcaa ggtaagtggg ttcattaccg
      301 cttatcaccg catattccag catgggcggc gaaaattatt gatgaggcct ggcgatgtga
      361 acaggaaaag gttcaggcga ttgtccgcaa cctggctcga caaaactgtt ccggggacag
      421 taagaacatt tgcagttaaa aatttagcta aacacacatg aattttcaga tgtgttttat
      481 ccgggtacta gagccaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt
      541 cgttttatct gttgtttgtc ggtgaacgct ctctactaga gtcacactgg ctcaccttcg
      601 ggtgggcctt tctgcgttta tatactagag ccaactcaaa attcacacct attaccttcc
      661 tctgcactta cacattcgtt aagtcatata tgtttttgac ttatccgctt cgaagagaga
      721 cactacctgc aatactagag attaaagagg agaaatacta gatgagtaca ggcatcgata
      781 aggacgtccg agagtgttgg ggcgtaactg cattatcagc gggtcatcaa attgcaatga
      841 atagcgcgtt tctggatatg gacttgctgt tgtgcgggga aaccggcacc ggcaaggaca
      901 cactggccaa ccgcattcac gagttgtcca gcaggtcggg accctttgtg ggcatgaact
      961 gcgccgccat tcccgagtcg ctggcagaga gccagttatt cggtgtggtc aacggtgcat
     1021 tcaccggcgt atgccgggct cgcgagggct acatagaggc ctccagtggt ggcaccttgt
     1081 acctggatga aatcgacagc atgccgttga gcctgcaagc caaactgctg cgtgtgttgg
     1141 agagtcgagg tatcgagcgt ctgggctcga ccgaatttat cccggtggat ctgcggatca
     1201 ttgcctcggc ccagcggcca ctggatgaac tggtggaaca aggacttttc cgtcgcgacc
     1261 tgttttttcg gctcaacgtg ctgacgcttc acttgccagc cttgcgcaaa cgtcgtgaac
     1321 agatcctgcc attgttcgac cagttcaccc agggtatcgc tgccgagttc ggacgtcccg
     1381 ctcctgcgct ggacagcggg cgtgtgcagc tgctgctcag ccacgactgg ccgggcaaca
     1441 tccgcgaatt gaagtctgcg gccaagcgct tcgtactcgg cttccccttg ctgggcgccg
     1501 accctgtgga agcgcttgac cctgccacgg ggctgcgcac gcaaatgcgc atcatcgaga
     1561 aaatgctcat ccaggatgcc ttgaagcggc acaggcacaa tttcgacgcg gtgcttcagg
     1621 agttggagtt gccaagacgc accctgtatc accgcatgaa ggaactggga gttgcagcgc
     1681 cgatcgctgc gacggccggg gtctaataaa acgcaggtgt ggttatgcta ctagagatta
     1741 aagaggagaa atactagatg agtcttgatg aaaggtttga ggatgatctg gacgaggagc
     1801 gggttccgaa tctggggata gttgccgaaa gtatttcgca actgggtatc gacgtgctgc
     1861 tatcgggtga gaccggcacg ggcaaagaca cgattgcccg acggattcat gagatgtcag
     1921 gccgcaaagg gcgcctggtg gcgatgaatt gcgcggccat tccggagtcc ctcgccgaga
     1981 gcgagttatt cggcgtggtc agcggtgcct acaccggcgc tgatcgctcc agagtcggtt
     2041 atgtcgaagc ggcgcagggc ggcacgctgt acctggatga gatcgatagc atgccgctga
     2101 gcctgcaagc caaattgctg agggtgctgg aaacccgagc gcttgaacgg ctgggttcga
     2161 cgtcgacgat caagctggat atctgcgtga tcgcctccgc ccaatgctcg ctggacgacg
     2221 ccgtcgagcg ggggcagttt cgtcgcgatc tgtattttcg cctgaacgtc ctgacactca
     2281 agcttcctcc gctacgtaac cagtctgatc gcatagttcc cctgttcaca cgttttacgg
     2341 ccgccgccgc gagggagctc ggtgttcccg ttcccgatgt ttgcccactg ctgcacaaag
     2401 tgctgctggg ccacgactgg cccggcaata tccgtgagct caaggcggca gccaaacgcc
     2461 atgtgctggg tttccccttg ctgggcgccg agccgcaggg cgaagagcac ttggcctgtg
     2521 ggctcaaatc gcaattgcga gtgatcgaaa aagccctgat tcaggagtcg ctcaagcgcc
     2581 acgacaattg tgtggattcg gtaagcctgg aactggacgt gccacgccgt acgctctatc
     2641 gacgcatcaa agaattgcag atctaataac caggcatcaa ataaaacgaa aggctcagtc
     2701 gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctct actagagtca
     2761 cactggctca ccttcgggtg ggcctttctg cgtttatata ctagaggccg gattatgtcc
     2821 gctgagtggg tcacggtccc ggatcagttc ccttgcgaag ctgaccgatg tttttgtgcc
     2881 aaaagctgtt gtggcaaaaa acggtttgcg caaagttttg tattacaaag aatttcacat
     2941 tttaaaatat ctttataaat caatcagtta tttctatttt caagctggca cggttattgc
     3001 tatagggctt gtactactag agattaaaga ggagaaatac tagatgcgta aaggagaaga
     3061 acttttcact ggagttgtcc caattcttgt tgaattagat ggtgatgtta atgggcacaa
     3121 attttctgtc agtggagagg gtgaaggtga tgcaacatac ggaaaactta cccttaaatt
     3181 tatttgcact actggaaaac tacctgttcc atggccaaca cttgtcacta ctttcggtta
     3241 tggtgttcaa tgctttgcga gatacccaga tcatatgaaa cagcatgact ttttcaagag
     3301 tgccatgccc gaaggttatg tacaggaaag aactatattt ttcaaagatg acgggaacta
     3361 caagacacgt gctgaagtca agtttgaagg tgataccctt gttaatagaa tcgagttaaa
     3421 aggtattgat tttaaagaag atggaaacat tcttggacac aaattggaat acaactataa
     3481 ctcacacaat gtatacatca tggcagacaa acaaaagaat ggaatcaaag ttaacttcaa
     3541 aattagacac aacattgaag atggaagcgt tcaactagca gaccattatc aacaaaatac
     3601 tccaattggc gatggccctg tccttttacc agacaaccat tacctgtcca cacaatctgc
     3661 cctttcgaaa gatcccaacg aaaagagaga ccacatggtc cttcttgagt ttgtaacagc
     3721 tgctgggatt acacatggca tggatgaact atacaaataa taatactaga gccaggcatc
     3781 aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg
     3841 tgaacgctct ctactagagt cacactggct caccttcggg tgggcctttc tgcgtttata
     3901 tactagtagc ggccgctgca ggcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     3961 ctgcggcgag cggtatcagc tcactcaaag gcggtaatct cgagtcccgt caagtcagcg
     4021 taatgctctg ccagtgttac aaccaattaa ccaattctga ttagaaaaac tcatcgagca
     4081 tcaaatgaaa ctgcaattta ttcatatcag gattatcgat accatatttt tgaaaaagcc
     4141 gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt
     4201 atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa
     4261 aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca
     4321 aaagcttatg catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa
     4381 aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata
     4441 cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca
     4501 ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg
     4561 ctgttttccc ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat
     4621 gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg
     4681 taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct
     4741 tcccatacaa tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat
     4801 acccatataa atcagcatcc atgttggaat ttaatcgcgg cctggagcaa gacgtttccc
     4861 gttgaatatg gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg
     4921 ttcatgatga tatattttta tcttgtgcaa tgtaacatca gagattttga gacacaacgt
     4981 ggctttgttg aataaatcga acttttgctg agttgaagga tcagatcacg catcttcccg
     5041 acaacgcaga ccgttccgtg gcaaagcaaa agttcaaaat caccaactgg tccacctaca
     5101 acaaagctct catcaaccgt ggctccctca ctttctggct ggatgatggg gcgattcagg
     5161 cctggtatga gtcagcaaca ccttcttcac gaggcagacc tcagcgctag cggagtgtat
     5221 actggcttac tatgttggca ctgatgaggg tgtcagtgaa gtgcttcatg tggcaggaga
     5281 aaaaaggctg caccggtgcg tcagcagaat atgtgataca ggatatattc cgcttcctcg
     5341 ctcactgact cgctacgctc ggtcgttcga ctgcggcgag cggaaatggc ttacgaacgg
     5401 ggcggagatt tcctggaaga tgccaggaag atacttaaca gggaagtgag agggccgcgg
     5461 caaagccgtt tttccatagg ctccgccccc tgacaagcat cacgaaatct gacgctcaaa
     5521 tcagtggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggcggctc
     5581 cctcgtgcgc tctcctgttc ctgcctttcg gtttaccggt gtcattccgc tgttatggcc
     5641 gcgtttgtct cattccacgc ctgacactca gttccgggta ggcagttcgc tccaagctgg
     5701 actgtatgca cgaacccccc gttcagtccg accgctgcgc cttatccggt aactatcgtc
     5761 ttgagtccaa cccggaaaga catgcaaaag caccactggc agcagccact ggtaattgat
     5821 ttagaggagt tagtcttgaa gtcatgcgcc ggttaaggct aaactgaaag gacaagtttt
     5881 ggtgactgcg ctcctccaag ccagttacct cggttcaaag agttggtagc tcagagaacc
     5941 ttcaaaaaac cgccctgcaa ggcggttttt tcgttttcag agcaagagat tacgcgcaga
     6001 ccaaaacgat ctcaagaaga tcatcttatt aaggggtctg acgctcagtg gaacgaaaac
     6061 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     6121 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     6181 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6241 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6301 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6361 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6421 tctattccat ggtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     6481 aaaataggcg tatcacgagg cagaatttca gataaaaaaa atccttagct ttcgctaagg
     6541 atgatttctg g
//
