变形链球菌gcp基因敲除菌株表达谱基因芯片

Streptococcus mutans gcp gene knockout strains expression profile gene chip

背景:前期研究中经证实变形链球菌内部存在单磷酸鸟苷环二聚体信号通路,构建了变形链球菌 gcp 基因敲除菌株。目的:比较变形链球菌野生菌种和 gcp 基因突变菌株基因表达的差异情况,筛选与生物膜相关的基因,进入后续研究。方法:提取两种细菌的总 RNA,反转录后分别用 cy3 和 cy5 染色。与基因芯片杂交后,扫描结果,进行数据分析,获取差异基因信息,对筛选的基因进行 Real-Time PCR 验证。结果与结论:差异基因主要与糖代谢、生物膜形成有关,选择了 2 个基因进行验证,PCR 结果与芯片结果相符合。变形链球菌 gcp 基因敲除后,突变菌株 ahpC 基因表达上调,磷酸转移酶系统基因表达下调,说明这 2个基因与 c-di-GMP 信号通路的下游途径相关。

BACKGROUND： Previous studies have confirmed the presence of bis-（3＇-5＇）-cyclic dimeric guanosine monophosphate signaling pathway in Streptococcus mutans, which construct the streptococcus mutans gcp gene knockout strains. OBJECTIVE： To compare the gene expression differences between Streptococcus mutans wild strains and gcp mutant strains, and to screen the biofilm-related genes from them for the follow-up study. METHODS： The total RNA of two kinds of strains were extracted and stained with cy3 and cy5 respectively after reverse transcription： The gene chip was scanned after hybridization and the differential gene were obtained through the data analysis. The different expression genes were verified by real-time PCR. RESULTS AND CONCLUSION： Differential genes were mainly relative about glucose metabolism and biofilm formation. We selected two genes for real-time PCR verification. The PCR results were consistent with the microarray results. After Streptococcus mutans gcp gene knockout, the gene expressions of gcp mutant strains were upregulated and the gene expressions of phosphotransferase system were downregulated, this result suggested that two different genes were related with the c-di-GMP signal pathway downstream.