海水希瓦氏菌(Shewanella aquimarina)PCR检测方法

doi:doi:10.3969/j.issn.1008-0864.2013.02.26

›› 2013, Vol. 15 ›› Issue (2): 168-172.DOI: doi:10.3969/j.issn.1008-0864.2013.02.26

海水希瓦氏菌(Shewanella aquimarina)PCR检测方法

王轶南,穆晓虎,封妮莎,丁君,常亚青*

(大连海洋大学, 农业部北方海水增养殖重点实验室, 辽宁大连, 116023)

收稿日期:2012-05-06 修回日期:2012-11-15 出版日期:2013-04-15 发布日期:2013-04-16
通讯作者: 常亚青,教授,博士生导师,主要从事海洋动物遗传育种和海水养殖研究。E-mail:yqchang@dlou.edu.cn
作者简介:王轶南,助理研究员,博士,主要从事海洋生物和水产动物病害研究。E-mail:wangyinan@dlou.edu.cn。
基金资助:
国家海洋局海洋公益项目（201105007)；辽宁省教育厅项目（L2012263）；国家863计划项目（2012AA10A412）；国家自然科学基金项目（30972269）；辽宁省科技计划项目（2007203004）资助。

PCR Detection Method of Shewanella aquimarina

WANG Yi-nan, MU Xiao-hu, FENG Ni-sha, DING Jun, CHANG Ya-qing*

(Key Laboratory of Mariculture &|Stock Enhancement in North Chinas Sea, Ministry of Agriculture, Dalian Ocean University, Dalian116023, Liaoning, China)

Received:2012-05-06 Revised:2012-11-15 Online:2013-04-15 Published:2013-04-16

摘要/Abstract

摘要：

海水希瓦氏菌（Shewanella aquimarina）是虾夷马粪海胆的一种致病菌。为建立海水希瓦氏菌PCR快速检测方法，根据海水希瓦氏菌gyrB基因的高变区序列设计3对引物，随后进行了其特异性和灵敏度分析。结果显示引物SA-9可扩增出片段大小为815 bp的海水希瓦氏菌特异片段。引物可检测的最低细菌量为4.6×103 cfu/mL,最低DNA含量为3.15×10-4 ng/μL。表明以SA-9为引物的PCR检测方法特异性强，灵敏度高。另外以该方法对海胆及其生境样品进行初步检测，发现健康海胆、养殖海水及投喂海带为阴性，而自然海域的海水具有一定量的海水希瓦氏菌。

关键词: 海水希瓦氏菌;gyrB基因;PCR;特异性；灵敏度

Abstract:

Shewanella aquimarina is one of the pathogenic bacteria of sea urchin Strongylocentrotus intermedius. To establish PCR detection process of S. aquimarina, 3 pairs of primer were designed according to the specified region of gyrB genes of S. aquimarina, and the specificity and sensitivity were analyzed. The results showed that the amplified fragments of PCR applied with primer SA-9 was agreed with the expected sizes. SA-9 could detect the minimum bacteria concentration of 4.6×103 cfu/mL, and the minimum detectable concentration of bacteria DNA was 0.315 pg/μL. These experiments showed that primer SA-9 had high specificity and sensitivity to S. aquimarina. S. aquimarina in coelomic fluid of sea urchins and the environmental samples were detected by the method. The results of healthy sea urchins, breeding seawater and feeding kelp were negative, while the sample of seawater from natural sea area had a certain amount of S. aquimarina.

Key words: Shewanell aquimarina, gyrB gene, PCR, specification, sensitivity

中图分类号:

S917.3

王轶南,穆晓虎,封妮莎,丁君,常亚青*. 海水希瓦氏菌(Shewanella aquimarina)PCR检测方法[J]. , 2013, 15(2): 168-172.

WANG Yi-nan, MU Xiao-hu, FENG Ni-sha, DING Jun, CHANG Ya-qing*. PCR Detection Method of Shewanella aquimarina[J]. , 2013, 15(2): 168-172.

海水希瓦氏菌(Shewanella aquimarina)PCR检测方法

PCR Detection Method of Shewanella aquimarina

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