LOCUS       sequence-456754-        3562 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PMetE promoter driving expression of sfGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3562)
  AUTHORS   Piorino F, Johnson S, Styczynski MP
  TITLE     A Cell-Free Biosensor for Assessment of Hyperhomocysteinemia.
  JOURNAL   ACS Synth Biol. 2023 Aug 18;12(8):2487-2492. doi: 
            10.1021/acssynbio.3c00103. Epub 2023 Jul 17.
  PUBMED    37459448
REFERENCE   2  (bases 1 to 3562)
  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.3c00103"; journalName: "ACS Synth Biol"; date: 
            "2023-08-18- 18"; volume: "12"; issue: "8"; pages: "2487-2492"
COMMENT     Sequence Label: pMetEGFP
FEATURES             Location/Qualifiers
     source          1..3562
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      17..605
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     506..525
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     RBS             1759..1781
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2437..2484
                     /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             2485..2508
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(2563..2581)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2577..2624
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2642..3457)
                     /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     3365..3384
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3449..3468
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
       61 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
      121 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
      181 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
      241 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
      301 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
      361 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
      421 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
      481 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
      541 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
      601 ggaaaaacgc cagcaacgcg atcccgcgaa atttacaggc gcgctggtga tcctggcctc
      661 actgtgggtg catcgtaagt gggtcgctcc gcgctcttca caaacggcag atgatcgcag
      721 gcgtgattgc gcgctgagcg aataaacgct tccgtaactg gctgacgctg ctcgccatcg
      781 cgcacggcgg cgtacagtcg gctccacaag ccttcgccca gggttttggt caccaccaga
      841 ccctggcgct caaaactctc tactacccaa tgcggtagcg cggcaatacc catccgcgcg
      901 gcaaccatct gaatcaacaa taaggtgtta tcgacgcttt tcagtgacgg gctgacgcct
      961 gccggctgaa gaaaatgccg ccagacatcc agtcgactac gctgcaccgg ataaattaat
     1021 agcgtctcgc tggcgagatc ttccggtgta attcgcgttt tcgccgccag tggatggtca
     1081 ggtgctaaca ccagacgcac ttcatagtcg aacatcggcg aataatgcag gccactgcgc
     1141 ggcagaatat cggacgtcat taccagatcc agctctccct gttgcaaggc gggctgcggg
     1201 tcaaatgtca cgcccgattt aaaatccatc tctacctgcg gccagttctt atggaaattt
     1261 tctaacgcgg gtgtcagcca ctgaatacag ctatggcact caatggcaat gcgcagacgc
     1321 gtctgctgcg gttcattgca ggcttgcagg gcctggctaa tttgcggcag tacctggttt
     1381 gccagttgca acaggatttc tccctgcggt gtaaagcgta gcggctggct cttacgcaca
     1441 aatagccgga agccaaggcg ttgttccaga tcgctaaact ggtgagacag ggcggattgc
     1501 gtctgatgca acgtcgccgc agcggctgcg agcgagccgc agttccgcaa cgcttgtagc
     1561 gttttcaggt gttttacttc gatcatgaaa gtccttcact tcggcatgaa taatttgcgc
     1621 ttgaggaata tacagtaacc gccaattatg gatgtgtaaa catctggacg gctaaaatcc
     1681 ttcgtctttt aaatttatgg tgcgttggct gcgtttctcc accccggtca cccacaacgg
     1741 tttccctcta gaaataattt tgtttaactt taagaaggag atatacatat gagcaaaggt
     1801 gaagaactgt ttaccggcgt tgtgccgatt ctggtggaac tggatggcga tgtgaacggt
     1861 cacaaattca gcgtgcgtgg tgaaggtgaa ggcgatgcca cgattggcaa actgacgctg
     1921 aaatttatct gcaccaccgg caaactgccg gtgccgtggc cgacgctggt gaccaccctg
     1981 acctatggcg ttcagtgttt tagtcgctat ccggatcaca tgaaacgtca cgatttcttt
     2041 aaatctgcaa tgccggaagg ctatgtgcag gaacgtacga ttagctttaa agatgatggc
     2101 aaatataaaa cgcgcgccgt tgtgaaattt gaaggcgata ccctggtgaa ccgcattgaa
     2161 ctgaaaggca cggattttaa agaagatggc aatatcctgg gccataaact ggaatacaac
     2221 tttaatagcc ataatgttta tattacggcg gataaacaga aaaatggcat caaagcgaat
     2281 tttaccgttc gccataacgt tgaagatggc agtgtgcagc tggcagatca ttatcagcag
     2341 aataccccga ttggtgatgg tccggtgctg ctgccggata atcattatct gagcacgcag
     2401 accgttctgt ctaaagatcc gaacgaaaaa ggcacgcggg accacatggt tctgcacgaa
     2461 tatgtgaatg cggcaggtat tacgtggagc catccgcagt tcgaaaaata agtcgaccgg
     2521 ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag
     2581 cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa gccaattctg
     2641 attagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa
     2701 taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc
     2761 ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca tcaatacaac
     2821 ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga
     2881 ctgaatccgg tgagaatggc aaaagcttat gcatttcttt ccagacttgt tcaacaggcc
     2941 agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt
     3001 gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa acaggaatcg
     3061 aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat
     3121 attcttctaa tacctggaat gctgttttcc cggggatcgc agtggtgagt aaccatgcat
     3181 catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt
     3241 ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa
     3301 acaactctgg cgcatcgggc ttcccataca atcgatagat tgtcgcacct gattgcccga
     3361 cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg
     3421 gcttcgagca agacgtttcc cgttgaatat ggctcataac accccttgta ttactgttta
     3481 tgtaagcaga cagttttatt gttcatgatg atatattttt atcttgtgca atgtaacatc
     3541 agagattttg agacacaacg tg
//
