Quantitative Real-Time PCR (qRT-PCR)

SSA-1575, SSA-2379, SSA-0016, and SSA-0391, randomly selected DEGs, were performed qRT-PCR to validate the results of RNA-seq. cDNA was synthesized from 2 μg of RNA using the SuperScript II reverse transcriptase (Invitrogen) followed the manufacturer's recommendation. The primer sequences are listed in Table 1. The gyrA gene was used as reference gene for calculation of the relative target gene expression using the 2^−ΔΔCt method. All qPCR results are presented as ratios of the ΔCcpA/wild type levels relative to gyrA transcripts. The experiments were repeated three times independently, and three replicates were involved in each sample. Data were analyzed with GraphPad Prism (version 5.0) software using Student's t-test (P-value below 0.05 was considered statistical significance).

List of the primer sequences used in qPCR analysis.