LOCUS       sequence-331025-        3909 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  cell-free cadmium sensing reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3909)
  TITLE     Sensitizing cell-free molecular detection using cascaded genetic 
            circuitry
REFERENCE   2  (bases 1 to 3909)
  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"
COMMENT     Sequence Label: pJBL7051
FEATURES             Location/Qualifiers
     source          1..3909
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(49..153)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     221..239
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(277..299)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     399..418
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     612..634
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     626..643
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     627..643
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             join(774..776,777..1490)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             774..1487
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(804..832)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1427..1447
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(1625..1642)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1795..1841
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(1937..1953)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1937..1953)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1950..1972)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1961..1977
                     /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)."
     promoter        complement(join(1985..1991,1992..2009,2010..2015))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2030..2051
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2168..2185)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2338..2926)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2418..2437)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3097..3888,3889..3909,1..48))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3720..3739
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
       61 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
      121 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
      181 cgtctaagaa accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc
      241 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
      301 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
      361 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
      421 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
      481 caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
      541 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
      601 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtaagatct cggctgaatt
      661 ctaaaagatc tataacttga ctctgtagtt gctacagggt gtgcaatcgg ttacgtcgac
      721 tctcgagtga gattgttgac ggtaccgtat tttggatcta ggaggaagga tctatgagca
      781 aaggagaaga acttttcact ggagttgtcc caattcttgt tgaattagat ggtgatgtta
      841 atgggcacaa attttctgtc cgtggagagg gtgaaggtga tgctacaaac ggaaaactca
      901 cccttaaatt tatttgcact actggaaaac tacctgttcc gtggccaaca cttgtcacta
      961 ctctgaccta tggtgttcaa tgcttttccc gttatccgga tcacatgaaa cggcatgact
     1021 ttttcaagag tgcaatgccc gaaggttatg tacaggaacg cactatatct ttcaaagatg
     1081 acgggaccta caagacgcgt gctgaagtca agtttgaagg tgataccctt gttaatcgta
     1141 tcgagttaaa gggtattgat tttaaagaag atggaaacat tcttggacac aaactcgagt
     1201 acaactttaa ctcacacaat gtatacatca cggcagacaa acaaaagaat ggaatcaaag
     1261 ctaacttcaa aattcgccac aacgttgaag atggttccgt tcaactagca gaccattatc
     1321 aacaaaatac tccaattggc gatggccctg tccttttacc agacaaccat tacctgtcga
     1381 cacaatctgt cctttcgaaa gatcccaacg aaaagcgtga ccacatggtc cttcttgagt
     1441 ttgtaactgc tgctgggatt acacatggca tggatgagct ctacaaataa ggatctgaag
     1501 cttgggcccg aaaaaaaact catctcagaa gaggatctga atagcgccgt cgaccatcat
     1561 catcatcatt gagtttaaac ggtctccagc ttggctgttt tggcggatga gagaagattt
     1621 tcagcctgat acagattaaa tcagaacgca gaagcggtct gataaaacag aatttgcctg
     1681 gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta
     1741 gcgccgatgg tagtgtgggg tctccccatg cgagagtagg gaactgccag gcatcaaata
     1801 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     1861 tctgaaagga ggaactatat ccggatcgag atcctctaga gtcgacctgc aggcatgcaa
     1921 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     1981 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     2041 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     2101 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     2161 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     2221 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     2281 tgtgagcaaa aggccagcaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     2341 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     2401 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     2461 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     2521 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     2581 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     2641 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     2701 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     2761 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     2821 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     2881 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     2941 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     3001 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     3061 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     3121 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     3181 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     3241 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     3301 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     3361 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     3421 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     3481 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     3541 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     3601 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     3661 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     3721 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     3781 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     3841 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     3901 aaggcaaaa
//
