LOCUS       sequence-341118-        2563 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  cell-free lac sensor with GFP output.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2563)
  AUTHORS   Jung JK, Rasor BJ, Rybnicky GA, Silverman AD, Standeven J, Kuhn R, 
            Granito T, Ekas HM, Wang BM, Karim AS, Lucks JB, Jewett MC
  TITLE     At-Home, Cell-Free Synthetic Biology Education Modules for 
            Transcriptional Regulation and Environmental Water Quality 
            Monitoring.
  JOURNAL   ACS Synth Biol. 2023 Oct 20;12(10):2909-2921. doi: 
            10.1021/acssynbio.3c00223. Epub 2023 Sep 12.
  PUBMED    37699423
REFERENCE   2  (bases 1 to 2563)
  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.3c00223"; journalName: "ACS Synth Biol"; date: 
            "2023-10-20- 20"; volume: "12"; issue: "10"; pages: "2909-2921"
COMMENT     Sequence Label: pJBL7079
FEATURES             Location/Qualifiers
     source          1..2563
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     192..211
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        join(341..346,347..364,365..371)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    379..395
                     /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)."
     primer_bind     384..406
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     RBS             446..468
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1124..1171
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     CDS             1172..1195
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(1250..1268)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1264..1311
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1329..2144)
                     /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="MSHIQRETSCSKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2052..2071
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2136..2155
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      join(2266..2563,1..291)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
       61 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
      121 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
      181 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
      241 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
      301 aacgcgatcc cgcgaaattc actcattagg caccccaggc tttacacttt atgcttccgg
      361 ctcgtataat gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctgagacc
      421 acaacggttt ccctctagaa ataattttgt ttaactttaa gaaggagata tacatatgag
      481 caaaggtgaa gaactgttta ccggcgttgt gccgattctg gtggaactgg atggcgatgt
      541 gaacggtcac aaattcagcg tgcgtggtga aggtgaaggc gatgccacga ttggcaaact
      601 gacgctgaaa tttatctgca ccaccggcaa actgccggtg ccgtggccga cgctggtgac
      661 caccctgacc tatggcgttc agtgttttag tcgctatccg gatcacatga aacgtcacga
      721 tttctttaaa tctgcaatgc cggaaggcta tgtgcaggaa cgtacgatta gctttaaaga
      781 tgatggcaaa tataaaacgc gcgccgttgt gaaatttgaa ggcgataccc tggtgaaccg
      841 cattgaactg aaaggcacgg attttaaaga agatggcaat atcctgggcc ataaactgga
      901 atacaacttt aatagccata atgtttatat tacggcggat aaacagaaaa atggcatcaa
      961 agcgaatttt accgttcgcc ataacgttga agatggcagt gtgcagctgg cagatcatta
     1021 tcagcagaat accccgattg gtgatggtcc ggtgctgctg ccggataatc attatctgag
     1081 cacgcagacc gttctgtcta aagatccgaa cgaaaaaggc acgcgggacc acatggttct
     1141 gcacgaatat gtgaatgcgg caggtattac gtggagccat ccgcagttcg aaaaataagt
     1201 cgaccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa
     1261 taactagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaagcc
     1321 aattctgatt agaaaaactc atcgagcatc aaatgaaact gcaatttatt catatcagga
     1381 ttatcaatac catatttttg aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg
     1441 cagttccata ggatggcaag atcctggtat cggtctgcga ttccgactcg tccaacatca
     1501 atacaaccta ttaatttccc ctcgtcaaaa ataaggttat caagtgagaa atcaccatga
     1561 gtgacgactg aatccggtga gaatggcaaa agcttatgca tttctttcca gacttgttca
     1621 acaggccagc cattacgctc gtcatcaaaa tcactcgcat caaccaaacc gttattcatt
     1681 cgtgattgcg cctgagcgag acgaaatacg cgatcgctgt taaaaggaca attacaaaca
     1741 ggaatcgaat gcaaccggcg caggaacact gccagcgcat caacaatatt ttcacctgaa
     1801 tcaggatatt cttctaatac ctggaatgct gttttcccgg ggatcgcagt ggtgagtaac
     1861 catgcatcat caggagtacg gataaaatgc ttgatggtcg gaagaggcat aaattccgtc
     1921 agccagttta gtctgaccat ctcatctgta acatcattgg caacgctacc tttgccatgt
     1981 ttcagaaaca actctggcgc atcgggcttc ccatacaatc gatagattgt cgcacctgat
     2041 tgcccgacat tatcgcgagc ccatttatac ccatataaat cagcatccat gttggaattt
     2101 aatcgcggct tcgagcaaga cgtttcccgt tgaatatggc tcataacacc ccttgtatta
     2161 ctgtttatgt aagcagacag ttttattgtt catgatgata tatttttatc ttgtgcaatg
     2221 taacatcaga gattttgaga cacaacgtga gatcaaagga tcttcttgag atcctttttt
     2281 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     2341 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     2401 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     2461 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     2521 gtcgtgtctt accgggttgg actcaagacg atagttaccg gat
//
