溃疡性结肠炎相关肠道菌群荧光定量PCR检测方法的建立

大理大学学报 ›› 2019, Vol. 4 ›› Issue (2): 33-.

溃疡性结肠炎相关肠道菌群荧光定量PCR检测方法的建立

段晓梅，李超男，耿福能，刘彬，赵昱，李玥

（1.云南省昆虫生物医药研发重点实验室，云南大理671000；2.药用特种昆虫开发国家地方联合工程研究中心，
云南大理671000；3.中国西南药用昆虫及蛛形类资源开发利用协同创新中心，云南大理671000；
4.药用美洲大蠊四川省重点实验室，成都610000）

收稿日期:2018-10-31 修回日期:2018-12-23 出版日期:2019-02-15 发布日期:2019-02-15
通讯作者: 李玥，研究员实习员，E-mail：liyue0208@163.com.
作者简介:段晓梅，硕士研究生，主要从事中药抗炎药理学研究.
基金资助:
国家自然科学基金资助项目（81660605；81860742；81860765）：云南省地方本科高校（部分）基础研究联合专
项资助项目（2017FH001-108）；云南省教育厅科学研究基金产业化培育项目（2016CYH15）；药用特种昆虫开
发国家地方联合工程研究中心创新能力建设项目（云发改高技〔2015〕395号）；云南省昆虫生物医药研发重点
实验室专项经费资助项目（2015DG030）；云南省2011协同创新中心资助项目（云教科〔2012〕25号）

Establishment of a Real-time PCR Method for Detection of Gut Bacteria in Patients with Ulcerative Colitis

Duan Xiaomei, Li Chaonan, Geng Funeng, Liu Bin, Zhao Yu, Li Yue

（1. Yunnan Provincial Key Laboratory of Entomological Biopharmaceutical R&D, Dali, Yunnan 671000, China；
2. National-Local Joint Engineering Research Center of Entomoceutics, Dali, Yunnan 671000, China；3.Yunnan Provincial
Collaborative Innovation Center for Entomoceutics, Dali , Yunnan 671000, China; 4. Key Laboratory of
Periplaneta americana in Sichuan Province, Chengdu 610000, China）

Received:2018-10-31 Revised:2018-12-23 Online:2019-02-15 Published:2019-02-15

摘要/Abstract

摘要： ［摘要］目的：为了进一步探索菌群失调对溃疡性结肠炎（Ulcerative Colitis，UC）的影响，建立与UC相关的肠道菌群荧光定量
qRT-PCR检测方法。方法：通过筛选确定7种与UC发病及病程进展有较强相关性的肠道微生物种类，设计PCR引物，扩增出
标志性片段重组到质粒中作为标准品，利用实时荧光定量PCR技术建立相关肠道菌的定量检测方法。结果：成功建立了检测
双歧杆菌属等优势菌群、大肠埃希菌等条件致病菌及肠道总菌群的qRT-PCR方法，各相关菌种重组质粒建立的标准曲线相关
系数≥0.998。结论：通过实验验证筛选了7种与UC相关的肠道菌群，并建立其绝对定量的qRT-PCR检定方法，具有特异性及
准确度高等特点。

关键词: ［关键词］荧光定量PCR, 肠道菌群, 溃疡性结肠炎, 重组质粒

Abstract: 〔Abstract〕Objective: To establish a quantitative real-time PCR method for detecting gut bacteria in patients with ulcerative colitis
in order to further explore the effects of imbalanced gut flora on ulcerative colitis（UC）. Methods: We identified 7 gut microbes that
were strongly associated with the pathogenesis of UC. Specific PCR primers were designed to construct a standard recombinant plasmid
containing specific amplified fragments. The absolute quantitative method of real-time PCR was established by using recombinant
plasmids as standards. Results: The real-time PCR method for the detection of such target bacteria as Bifidobacterium, Escherichia
coli, and total intestinal flora, was successfully established. The standard curve of gut bacteria showed a good linear relation with
correlation coefficient ≥0.998. Conclusion: The SYBR Green real-time PCR method can detect seven intestinal bacteria in ulcerative
colitis specifically and provide a convenient and effective method for exploring the correlation between the imbalance of gut bacteria
and UC pathogenesis.

Key words: 〔Key words〕Real-time PCR, gut bacteria, Ulcerative Colitis recombinant plasmid

段晓梅，李超男，耿福能，刘彬，赵昱，李玥. 溃疡性结肠炎相关肠道菌群荧光定量PCR检测方法的建立[J]. 大理大学学报, 2019, 4(2): 33-.

Duan Xiaomei, Li Chaonan, Geng Funeng, Liu Bin, Zhao Yu, Li Yue. Establishment of a Real-time PCR Method for Detection of Gut Bacteria in Patients with Ulcerative Colitis[J]. Journal of Dali University, 2019, 4(2): 33-.

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