LAMP检测无乳链球菌方法的建立及应用

doi:10.7668/hbnxb.2009.05.050

华北农学报 ›› 2009, Vol. 24 ›› Issue (5): 234-238. doi: 10.7668/hbnxb.2009.05.050

• 论文 • 上一篇

LAMP检测无乳链球菌方法的建立及应用

王永¹, 赵新¹, 景海春², 兰青阔¹, 朱珠¹, 程奕⁵

1. 天津市农业科学院中心实验室, 天津 300381;
2. Plant Pathogen Interactions Division,Rothamsted Research,AL5 2JQ UK

收稿日期:2009-03-25 出版日期:2009-10-28
通讯作者: 程奕(1963-)，女，安徽歙县人，研究员，硕士，主要从事农产品质量检测技术研究。
作者简介:王永(1977-)，男，天津人，助理岩究员，硕士，主要从事农产品安全质量分子检测技术研究。
基金资助:
科技部国际合作项目(2006DFA32380);天津市农业科学院院偿基金项目(05011)

Development and Application of Loop-Mediated Isothermal Amplification for Detection of Streptococcus agalactiae

WANG Yong¹, ZHAO Xin¹, JING Hai-chun², LAN Qing-kuo¹, ZHU Zhu¹, CHENG Yi⁵

1. Central Lab of Tianjin Academy of Agriculture Sciences, Tianjin 300381, China;
2. Plant Pathogen Interactions Division, Rothamsted Research, AL5 2JQ, UK

Received:2009-03-25 Published:2009-10-28

摘要/Abstract

摘要： 以无乳链球菌纤连蛋白fbs基因为主要研究对象，采取环介导等温扩增技术(Loop-Mediated Isothermal Amplification，LAMP)，针对fbs基因的6个区域设计4条特异性引物，利用一种链置换DNA聚合酶(Bst DNA polymerase)在63℃保温1h，通过荧光显色即可完成对无乳链球菌的检测工作。结果显示，LAMP方法能够特异性检测fbs基因，其检测灵敏度是常规PCR方法的100倍，并与实时荧光定量PCR方法相当。所建立的针对无乳链球菌fbs基因的LAMP检测方法具有高度的特异性及稳定性，结果可靠，非常适合无乳链球菌的快速检测。

关键词: 环介导等温扩增技术, 无乳链球菌, fbsB, 检测

Abstract: To develop a technique for detecting the fbsB gene of Streptococcus agalactiae using a novel DNA amplification method designated Loop-Mediated Isothermal Amplification (LAMP).The detection assay is based on the loop-mediated isothermal amplification (LAMP) reaction.The fbsB gene was amplified by a set of four specially primers that recognize six distinct sequences of the target.The amplification can be obtained in 1h by incubating all of the reagents in a single tube with Bst DNA polymerase at 63℃.Results showed that the developed LAMP assay demonstrated an exceptionally higher than the conventional PCR.This assay results was found to be 100 times more sensitive than the PCR assay,and consistent with the results of real-time PCR.Our results clearly demonstrate that the LAMP-based assay is a sensitive and reliable means for the detection of Streptococcus agalactiae.

Key words: Loop-mediated isothermal amplification (LAMP), Streptococcus agalactiae, fbsB, Detection

中图分类号:

S852.61

王永, 赵新, 景海春, 兰青阔, 朱珠, 程奕. LAMP检测无乳链球菌方法的建立及应用[J]. 华北农学报, 2009, 24(5): 234-238. doi: 10.7668/hbnxb.2009.05.050.

WANG Yong, ZHAO Xin, JING Hai-chun, LAN Qing-kuo, ZHU Zhu, CHENG Yi. Development and Application of Loop-Mediated Isothermal Amplification for Detection of Streptococcus agalactiae[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2009, 24(5): 234-238. doi: 10.7668/hbnxb.2009.05.050.

http://www.hbnxb.net/CN/Y2009/V24/I5/234

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