LOCUS       sequence-331028-        3907 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  cell-free copper sensing reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3907)
  TITLE     Sensitizing cell-free molecular detection using cascaded genetic 
            circuitry
REFERENCE   2  (bases 1 to 3907)
  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: pJBL7054
FEATURES             Location/Qualifiers
     source          1..3907
                     /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(772..774,775..1488)
                     /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             772..1485
                     /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(802..830)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1425..1445
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             1508..1537
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(1623..1640)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1793..1839
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(1935..1951)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1935..1951)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1948..1970)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1959..1975
                     /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(1983..1989,1990..2007,2008..2013))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2028..2049
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2166..2183)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2336..2924)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2416..2435)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3095..3886,3887..3907,1..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     3718..3737
                     /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 tttcttgacc ttccccttgc tggaaggttt atcctcggtt acgtcgactc
      721 tcgagtgaga ttgttgacgg taccgtattt tggatctagg aggaaggatc tatgagcaaa
      781 ggagaagaac ttttcactgg agttgtccca attcttgttg aattagatgg tgatgttaat
      841 gggcacaaat tttctgtccg tggagagggt gaaggtgatg ctacaaacgg aaaactcacc
      901 cttaaattta tttgcactac tggaaaacta cctgttccgt ggccaacact tgtcactact
      961 ctgacctatg gtgttcaatg cttttcccgt tatccggatc acatgaaacg gcatgacttt
     1021 ttcaagagtg caatgcccga aggttatgta caggaacgca ctatatcttt caaagatgac
     1081 gggacctaca agacgcgtgc tgaagtcaag tttgaaggtg atacccttgt taatcgtatc
     1141 gagttaaagg gtattgattt taaagaagat ggaaacattc ttggacacaa actcgagtac
     1201 aactttaact cacacaatgt atacatcacg gcagacaaac aaaagaatgg aatcaaagct
     1261 aacttcaaaa ttcgccacaa cgttgaagat ggttccgttc aactagcaga ccattatcaa
     1321 caaaatactc caattggcga tggccctgtc cttttaccag acaaccatta cctgtcgaca
     1381 caatctgtcc tttcgaaaga tcccaacgaa aagcgtgacc acatggtcct tcttgagttt
     1441 gtaactgctg ctgggattac acatggcatg gatgagctct acaaataagg atctgaagct
     1501 tgggcccgaa caaaaactca tctcagaaga ggatctgaat agcgccgtcg accatcatca
     1561 tcatcattga gtttaaacgg tctccagctt ggctgttttg gcggatgaga gaagattttc
     1621 agcctgatac agattaaatc agaacgcaga agcggtctga taaaacagaa tttgcctggc
     1681 ggcagtagcg cggtggtccc acctgacccc atgccgaact cagaagtgaa acgcagtagc
     1741 gccgatggta gtgtggggtc tccccatgcg agagtaggga actgccaggc atcaaataaa
     1801 acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt cggtgaactc
     1861 tgaaaggagg aactatatcc ggatcgagat cctctagagt cgacctgcag gcatgcaagc
     1921 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     1981 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     2041 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     2101 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     2161 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     2221 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     2281 tgagcaaaag gccagcaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     2341 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     2401 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     2461 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     2521 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     2581 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     2641 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     2701 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     2761 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     2821 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     2881 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     2941 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     3001 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     3061 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     3121 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     3181 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     3241 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     3301 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     3361 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     3421 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     3481 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     3541 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     3601 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     3661 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     3721 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     3781 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     3841 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     3901 ggcaaaa
//
