## 2 Results
### Expression Analysis of sRNAs and Target mRNAs under Different Acid Stress Conditions  
To preliminarily explore the biological function of sRNAs under acid stress condition, we detected the expression levels of _srn_884837 and _srn_133480, and their corresponding target genes (_glnQ_, _glnM_, _brpA_, and _relA_ for _srn_884837; _ffln_, _brpA_, and _relA_ for _srn_133480) related to acid tolerance under different acid stress conditions and at different time points. As shown in Figure 4A, we found that both sRNAs and their target genes can be induced during the first 0.5 h. The levels of target gene _ffln_ peaked at pH 4.5, and the target genes _brpA_, _relA_, _glnQ_, and _glnM_ presented higher expression levels at a pH of 6.5, 5.5, and 4.5, suggesting that _ffln_ has a major function in acid tolerance, particularly at pH 4.5, and that _brpA_, _relA_, _glnQ_, and _glnM_ play an extensive role in different acidic conditions (Figure 4B). Interestingly, the expression levels of _srn_884837 and _srn_133480 were lower at a pH of 5.5 and 4.5 compared with pH 7.5, whereas their target genes present relatively higher expression at pH 5.5 and 4.5 and lower expression at pH 7.5. The total relative expression levels of the two sRNAs and its target genes were shown in Figure 4C. The _p_-values for comparison of multiple means using Tukey honest significant difference (HSD) are shown in Table S3.  
Figure 4: _Cont_.  
Figure 3: Growth and vitality analysis of _S. mutans_ under different acid stress conditions. (**A**) Average absorbance (OD 600 nm) of _S. mutans_ grown at pH 7.5, 6.5, 5.5, and 4.5 for 0.5, 1, and 2 h. Cells can continue to grow at a pH of 7.5 and 6.5 from 0.5 to 2 h, and pH 5.5 at the first hour (_p_ \(<\) 0.05). The average absorbance value at 2 h was lower than those under pH 4.5 at 0.5 and 1 h (_p_ \(<\) 0.05); and (**B**) mean number of colony forming units (colony-forming unit (CFU) \(\times\) 10\({}^{10}\)) per mL at pH 7.5, 6.5, 5.5, and 4.5 for 0.5, 1, and 2 h. The survival bacteria at pH of 7.5, 6.5, and 5.5 increased over time and the survival bacteria at pH 4.5 decreased over time (_p_ \(<\) 0.05). The survival bacteria decreased at the acid was increased from 7.5 to 4.5 (_p_ \(<\) 0.001). The error bars indicate the standard deviations from triplicate experiments.  
Figure 4: qRT-PCR analysis of sRNAs (_srn_884837 and _srn_133480) and target mRNAs (_glnQ_, _glnM_, _ffh_, _brpA_, _relA_) under different acid stress conditions. (**A**) (**a**,**b**) present the expression levels of _srn_884837 and _srn_133480 in _S. mutant_, respectively. The expression level of each sRNA after growth at pH 7.5 for 0.5 h is defined as 1.0. The error bars display the standard deviations of three biological replicates. The two sRNAs present relatively lower expression at pH of 5.5 and 4.5 compared at pH 7.5 (_p_ < 0.05 for _srn_884837 and _srn_133480 at 0.5, 1, 2 h, respectively). The _p_-values for comparison of multiple means using Tukey’s honest significant difference (HSD) are detailed in Table S3; (**B**) (**a**–**e**) present the expression levels of the target genes _glnQ_, _glnM_, _ffh_, _brpA_, and _relA_, respectively. The expression level of each gene after growth at pH 7.5 for 0.5 h is defined as 1.0. The error bars display the standard deviations of three biological replicates. The target genes present relatively higher expression at pH 5.5 and 4.5 compared with pH 7.5 (_p_ < 0.05 for _glnQ_, _glnM_, _ffh_, _brpA_, and _relA_, respectively). The _p_-values for comparison of multiple means are detailed in Table S3; and (**C**) a heat map showing the total relative expression patterns of the two sRNAs and five target genes at pH values from 7.5 to 4.5 at each time point. The expression level of sm133480 after growth at pH 7.5 for 0.5 h is defined as 1.0.