食品科学 ›› 2013, Vol. 34 ›› Issue (10): 263-267.doi: 10.7506/spkx1002-6630-201310058

• 分析检测 • 上一篇    下一篇

REP-PCR及ERIC-PCR法对分离自海产品副溶血性弧菌分型分析

马月姣,孙晓红,赵 勇,卢 瑛,吴启华,潘迎捷   

  1. 1.上海海洋大学食品学院,上海水产品加工及贮藏工程技术研究中心,上海 201306;
    2.缅因大学食品科学与人类营养系,美国 欧洛诺 04469
  • 收稿日期:2012-02-20 修回日期:2013-04-07 出版日期:2013-05-25 发布日期:2013-05-07
  • 通讯作者: 孙晓红 E-mail:xhsun@shou.edu.cn

REP-PCR and ERIC-PCR Analysis for the Typing of Vibrio parahaemolyticus Isolated from Sea Products Marketed in Shanghai

MA Yue-jiao,SUN Xiao-hong,ZHAO Yong,LU Ying,Vivian Chi-Hua WU,PAN Ying-jie   

  1. 1. Aquatic Products Processing and Storage Engineering Technology Research Center in Shanghai, College of Food Science and Technology, Shanghai Ocean University, Shanghai 201306, China;
    2. Department of Food Science and Human Nutrition, the University of Maine, Orono 04469, USA
  • Received:2012-02-20 Revised:2013-04-07 Online:2013-05-25 Published:2013-05-07

摘要:

目的:采用细菌基因外重复回文序列扩增(REP-PCR)和肠道细菌基因间重复序列扩增(ERIC-PCR)两种方法对副溶血性弧菌2株标准株,13株实验室保存菌株和73株分离菌株共88株进行分子分型。方法:通过REP和ERICPCR指纹图谱扩增,利用NTsys-pc软件采用Dice系数对指纹图谱进行聚类结果分析。结果:所有供试菌株可通过此两种方法进行分型得到清晰的指纹图谱,并反映出副溶血性弧菌具有丰富的遗传多样性,REP-PCR扩增出5~9条分布在609~4200bp之间的条带,可将副溶血性弧菌分为5个群12类型,分辨指数达0.93,其中tdh+株在相似系数0.86时可聚类在第Ⅰ群;ERIC-PCR扩增出5~11条分布在400~7593bp之间的条带,将副溶血性弧菌分为7个群11个类型,分辨指数为0.94,其中tdh+株在相似系数0.76时可聚类在第Ⅰ群。结论:两种方法均显现出很好的分型能力,能够很好地将环境分离tdh+菌株和标准菌株聚类在一起,其中REP-PCR较ERIC-PCR重复性更好。

关键词: 副溶血性弧菌, tdh, 肠内细菌基因组间重复序列分析, 细菌基因外重复回文序列扩增, 分型

Abstract:

Objective: The enterobacterial repetitive intergenic consensus sequence-polymerase chain reaction (ERIC-PCR)
and repetitive extragenic palindromic element-polymerase chain reaction (REP-PCR) were used to analyze the types of 88
Vibrio parahaemolyticus including 2 standard strains, 13 laboratory strains and 73 isolated strains. Methods: REP and ERIC
primers were used to amplify the repeated sequences. NTsys-pc software was used to calculate the discriminative index of
REP- and ERIC-PCR with Dice coefficients. Results: All 88 tested V. parahamolyticus could be typed by REP- and ERICPCR
clearly and revealed abundant genetic diversity in V. parahamolyticus. According to the results of REP-PCR-amplified
5–9 bands within 609–4200 bp, V. parahamolyticus was typed into 5 groups and 12 types with discrimination index of 0.93,
and the tdh+ strains with resolving index of 0.86 could be classified in Group I. According to ERIC-PCR-amplified 5–11
bands within 400–7593 bp, V. parahamolyticus was typed into 7 groups and 11 types with discrimination index of 0.94, and
the tdh+ strains with discrimination index of 0.76 could be classified in group I. Conclusion: The isolated tdh+ strains are
clustered together with standard strains using the two methods. Although both methods exhibit good typing capability, REPPCR
is more repeatable and stable than ERIC-PCR.

Key words: Vibrio parahaemolyticus, tdh, enterobacterial repetitive intergenic consensus sequence-polymerase chain reaction (ERIC-PCR), repetitive extragenic palindromic-polymerase chain reaction (REP-PCR), typingg