Journal: Toxins
Article Title: BrnQ Branched-Chain Amino Acid Transporters Influence Toxin Production by, but Not Growth of, Clostridium perfringens Type A Strain ATCC3624.
doi: 10.3390/toxins17040187
Figure Lengend Snippet: Figure 5. A comparison of expression of the brnQ genes by wild-type ATCC3624, its isogenic brnQ null mutants, and complementing strains. ATCC3624, the single brnQ null mutants, or complementing strains were inoculated into TY broth and then incubated anaerobically for 2 h at 37 ◦C. Bacteria were collected and pelleted via centrifugation. Total RNA was extracted from the pellets, and cDNA was made for RT-qPCR analyses. Average CT values were normalized to the housekeeping recA gene, and the fold differences in expression were calculated using the comparative CT method (2−∆∆CT). The mean values from three independent experiments are shown. The error bars indicate the S.D. (A) RT- qPCR for the expression of brnQ2 or brnQ3 was performed using cDNA from the wild-type parent, the brnQ1 null mutant, and the complementing strain after their incubation in a TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. (B) RT-qPCR for the expression of brnQ1 or brnQ3 was performed using cDNA from the wild type-parent, the brnQ2 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type. (C) RT-qPCR for the expression of brnQ1 or brnQ2 was performed using cDNA from the wild-type parent, the brnQ3 null mutant, and the complementing strain after their incubation in TY medium for 2 h. The mean values from three independent experiments are shown. The error bars indicate the S.D. * p < 0.05 relative to the wild type.
Article Snippet: The current study used C. perfringens type A strain ATCC3624 (ATCC®, Manassas VA, USA) and E. coli DH5α competent cells [New England Biolabs (NEB), Ipswich, MA, USA].
Techniques: Comparison, Expressing, Incubation, Bacteria, Centrifugation, Quantitative RT-PCR, Mutagenesis