Journal: Scientific Reports
Article Title: The novel EHEC gene asa overlaps the TEGT transporter gene in antisense and is regulated by NaCl and growth phase
Figure Lengend Snippet: Regulated transcription of asa . ( A ) Promoter activity of the cloned putative promoter region (black letters in Fig. 4A ) was measured as fluorescence intensity of E. coli Top-10 + pProbe-NT:: asa P (OD 600 = 0.8) after growth in LB (grey), in LB + 450 mM NaCl (blue) or in LB + 10 mM L-arginine (orange); NC I: negative control using the terminator region of the gene EDL933_1236 (violet letters in Fig. 4A ), NC II: pProbe-NT without insert. The mean values and standard deviations of all three replicates are shown here. ( B ) RT-qPCR threshold cycles of asa normalized to the 16S rRNA gene (∆cq). Lower ∆cq values indicate higher amounts of mRNA and vice versa . EHEC was grown in LB medium (left columns) or LB + 450 mM NaCl (right columns). Approximately 10 8 cells were harvested for total RNA extraction either at exponential phase (OD 600 = 0.2–0.3, blue) or at late exponential phase (OD 600 = 0.7 – 0.8, orange). The RNA was reverse transcribed into cDNA and quantified using RT-qPCR. The mean values and standard deviations of all three replicates are shown here.
Article Snippet: List of bacteria and incubation conditions The following bacterial strains were used: E. coli O157:H7 strain EDL933 (Collection de l’Institute Pasteur: CIP 106327, GenBank accession number CP008957.1), E. coli O157:H7 strain Sakai (Weihenstephan strain collection WS 4518, GenBank accession number NC_002695.1), E. coli LF82 (kindly provided by R. Balfour Sartor, GenBank accession number NC_011993.1), E. coli TOP-10 (Invitrogen, Paisley, UK).
Techniques: Activity Assay, Clone Assay, Fluorescence, Negative Control, Quantitative RT-PCR, RNA Extraction