【摘要】 目的应用PCR-DGGE和rep-PCR技术对肝硬化大鼠肝移植后肠道菌群多样性进行研究,探讨肠道菌群多样性的变化,并比较这两种方法在菌群分析中的作用。方法首先建立CCL4诱导肝硬化大鼠的肝移植模型,收取对照组、肝硬化成模时、肝移植后7 d和肝移植后30 d的大鼠粪便,提取细菌基因组DNA,采用PCR-DGGE和rep-PCR[BOX-PCR,ERIC-PCR,ERIC2-PCR,(GTG)5-PCR,REP-PCR]进行DNA指纹图谱分析。结果PCR-DGGE可明确将正常大鼠、肝硬化大鼠、肝移植大鼠分为3个簇,并显示出肝硬化、肝移植大鼠肠道菌群多样性明显增多。rep-PCR技术也可将各组分开,其中ERIC-PCR、ERIC2-PCR及REP-PCR三者扩增条带各组差异有显著性,鉴别效果更好。结论应用基于16S rRNA基因和细菌基因组间重复序列的指纹图谱技术对肝硬化大鼠肝移植后肠道微生态的研究具有重要价值,可对临床肝硬化、肝移植患者肠道微生态制剂的使用起到指导作用。更多还原

【Abstract】 Objective To explore the diversity of intestinal microbiota in liver transplantation model with hepatic cirrhosis by PCR-DGGE and rep-PCR and compare the two methods with each other.Method Hepatic cirrhosis model was established in rats by subcutaneous injection of 40% carbon tetrachloride and liver transplantation model was made with two-cuff technique.Then the feces in the following groups were collected: control group(A),hepatic cirrhosis model group(B),7 days after liver transplantation(C) and 30 days after liver transplantation(D).After extraction of bacterial genomic DNA from feces,the diversity of intestinal microbiota were analyzed with PCR-DGGE and rep-PCR(BOX-PCR,ERIC-PCR,ERIC2-PCR,(GTG)5-PCR,REP-PCR).Result The bacterial composition and structure could be divided into three clusters among the four groups by PCR-DGGE distinctly.And we found that there were higher bacterial richness in hepatic cirrhosis group and liver transplantation groups.Among the five rep-PCR genomic fingerprinting methods,ERIC-PCR,ERIC2-PCR and REP-PCR could also discriminate against each other according to the molecular fingerprinting.Conclusion Both of the two molecular fingerprinting methods(PCR-DGGE analysis and rep-PCR technique) can be applied to analyze the dynamics of intestinal bacterial diversity in liver transplantation model with hepatic cirrhosis.更多还原