SYBR GreenⅠ实时荧光定量PCR法分析结直肠癌患者肠道菌群变化  

SYBR GreenⅠ Real-Time Fluorescence Quantitative PCR Analysis of Variation of Intestinal Microflora in Patients with Colorectal Cancer

作　　者：郭世奎[1] 包维民[1] 龚昆梅[1] 邵剑春[2] 陈弟[2] 王昆华[1] 

机构地区：[1]昆明医学院附属昆华医院(云南省第一人民医院)普外一科,昆明650032 [2]昆明市第一人民医院检验科,昆明650011

出　　处：《中国普外基础与临床杂志》2010年第5期463-468,共6页Chinese Journal of Bases and Clinics In General Surgery

基　　金：云南省应用基础研究基金资助(项目编号:2005C0074M;2008ZC085M);昆明医学院硕士研究生创新基金资助(项目编号:2009S02)~~

摘　　要：目的应用SYBR GreenⅠ实时荧光定量PCR法分析结直肠癌患者肠道菌群的变化,探讨肠道相关分子微生态在其发病中的作用及意义。方法根据细菌的16S rRNA基因序列设计双歧杆菌属、乳酸杆菌属和大肠杆菌的特异性引物,根据细菌的ddl基因序列设计粪肠球菌及屎肠球菌的特异性引物。收集结直肠癌患者术前粪便标本30份(结直肠癌组)及正常对照标本30份(正常对照组),提取细菌基因组DNA,应用SYBR GreenⅠ实时荧光定量PCR反应测定5种细菌的数量。结果正常对照组与结直肠癌组粪便中细菌数量分别为双歧杆菌属:9.25±0.83比4.52±0.49,乳酸杆菌属:7.45±0.37比5.46±0.12,结直肠癌组的数量明显减少(P<0.05);大肠杆菌:4.68±0.32比5.82±0.47,粪肠球菌:4.95±0.24比10.60±0.30,屎肠球菌:5.03±0.43比5.74±0.16,结直肠癌组的数量明显增多(P<0.05)。结论结直肠癌患者粪便中双歧杆菌及乳酸杆菌数量较正常对照明显减少,而大肠杆菌,粪肠球菌和屎肠球菌的数量较正常对照明显增多,提示结直肠癌的发生、发展与肠道菌群有一定关系。Objective To analyze the variation of intestinal microflora in patients with colorectal cancer by SYBR GreenⅠreal-time fluorescence quantitative PCR and reveal the role and significance of intestinal microflora in the colorectal cancer-associated molecular pathogenesis.Methods A set of 16S rRNA gene group of species-specific primers for Bifidobacterium spp.,Lactobacillus group,Escherichia coli,and ddl gene-targeted species-specific primers for Enterococcus faecalis and feces Enterococcus were designed.Patients with colorectal cancer（colorectal cancer group,n=30） and healthy volunteers（normal control group,n=30） were included and whose feces were collected to extract bacterial genome DNA.SYBR GreenⅠ real-time fluorescence quantitative PCR was used to analyze the five mentioned bacterial amounts.Results Level of Bifidobacterium spp.（4.52±0.49） and Lactobacillus group（5.46±0.12） in colorectal cancer group were significantly lower than those（9.25±0.83 and 7.45±0.37） of normal control group（P〈0.05）,whereas levels of Escherichia coli（5.82±0.47）,Enterococcus faecalis（10.6±0.30） and feces Enterococcus（5.74±0.16） in colorectal cancer group were significantly higher than those（4.68±0.32,4.95±0.24,and 5.03±0.43） of normal control group（P〈0.05）.Conclusions The fecal microflora composition of patients with colorectal cancer is significantly decreased in Bifidobacterium spp.and Lactobacillus group,whereas increased in Escherichia coli,Enterococcus faecalis,and feces Enterococcus.These data underline that the occurrence and progress of colorectal cancer may be related to intestinal microflora.

关 键 词：结直肠癌 肠道菌群 实时荧光定量PCR SYBR GreenⅠ 

分 类 号：R735.34[医药卫生—肿瘤]