LOCUS       sequence-194044-        1398 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Tetracycline inducible expression of GFPmut2 using autoregulation; 
            weak ribosome binding site; GFPmut2 ORF can be replaced with gene of
            interest by isothermal assembly.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1398)
  AUTHORS   Hensel Z
  TITLE     A plasmid-based Escherichia coli gene expression system with 
            cell-to-cell variation below the extrinsic noise limit.
  JOURNAL   PLoS One. 2017 Oct 30;12(10):e0187259. doi: 
            10.1371/journal.pone.0187259. eCollection 2017.
  PUBMED    29084263
REFERENCE   2  (bases 1 to 1398)
  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"; journalName: "PLoS One.";
            date: "2017-10-30"; volume: "12(10):e0187259. doi"; pages: " 
            10.1371/journal.pone.0187259. eCollection 2017"
COMMENT     Sequence Label: pZH518
FEATURES             Location/Qualifiers
     source          1..1398
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     23..42
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        complement(join(130..155,156..161,162..178,179..184,
                     185..203))
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    160..178
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    185..203
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     terminator      204..247
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(260..973)
                     /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="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFAYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(300..320)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     915..943
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     promoter        complement(join(1003..1028,1029..1034,1035..1051,1052..1057,
                     1058..1076))
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    1033..1051
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1058..1076
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     complement(1095..1114)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gctgctctac acctagcttc tgggcgagtt tacgggttgt taaaccttcg attccgacct
       61 cattaagcag ctctaatgcg ctgttaatca ctttactttt atctaatcta gacatggtga
      121 aatcctcctg gtcagtgcgt cctgctgatg tgctcagtat ctctatcact gatagggatg
      181 tcaatctcta tcactgatag ggaataaaac gaaaggccca gtctttcgac tgagcctttc
      241 gttttatgct agattattat ttgtatagtt catccatgcc atgtgtaatc ccagcagctg
      301 ttacaaactc aagaaggacc atgtggtctc tcttttcgtt gggatctttc gaaagggcag
      361 attgtgtgga caggtaatgg ttgtctggta aaaggacagg gccatcgcca attggagtat
      421 tttgttgata atggtctgct agttgaacgc ttccatcttc aatgttgtgt ctaattttga
      481 agttaacttt gattccattc ttttgtttgt ctgccatgat gtatacattg tgtgagttat
      541 agttgtattc caatttgtgt ccaagaatgt ttccatcttc tttaaaatca atacctttta
      601 actcgattct attaacaagg gtatcacctt caaacttgac ttcagcacgt gtcttgtagt
      661 tcccgtcatc tttgaaaaat atagttcttt cctgtacata accttcgggc atggcactct
      721 tgaaaaagtc atgctgtttc atatgatctg ggtatctcgc aaagcattga agaccatacg
      781 cgaaagtagt gacaagtgtt ggccatggaa caggtagttt tccagtagtg caaataaatt
      841 taagggtaag ttttccgtat gttgcatcac cttcaccctc tccactgaca gaaaatttgt
      901 gcccattaac atcaccatct aattcaacaa gaattgggac aactccagtg aaaagttctt
      961 ctcctttact catgcggtac ctttctcgaa tttaatgaat tcggtcagtg cgtcctgctg
     1021 atgtgctcag tatctctatc actgataggg atgtcaatct ctatcactga tagggacata
     1081 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     1141 ccaaaacccn ttgaagggcc ctgaangggc ttgtctgctc ccggcatccg cttacagaaa
     1201 agctgtgacc gtctccggga actgcnnggn tcaaaagttt tcaccgtcnt ccccgaaacc
     1261 gccgaaaaca aaagggcctc gggaaacncc tatttttaaa gggtaaatgg cnnnnaaant
     1321 aagggtttcn naaaagcccg gggggccctt tttcgnnnaa aatgtgcccn naaccccntt
     1381 tnnnttatnt tnccnaaa
//
