›› 2015, Vol. 58 ›› Issue (4): 460-466.

• 简报 • 上一篇    下一篇

应用种特异性PCR技术快速鉴定辣椒实蝇

黄振1, 陈韶萍2,3, 谢婧1, 郭琼霞2,*   

  1. (1. 福州机场出入境检验检疫局, 福州350209; 2. 福建出入境检验检疫局检验检疫技术中心, 福州350001;  3. 福建农林大学植物保护学院, 福州350002)
  • 出版日期:2015-04-20 发布日期:2015-04-20
  • 作者简介:黄振, 男, 1985年生, 福建莆田人, 博士研究生, 农艺师, 研究方向为农业昆虫与害虫防治, E-mail: 102768560@qq.com

Rapid identification of Bactrocera latifrons (Diptera: Tephritidae) by using species-specific PCR technique

HUANG Zhen1, CHEN Shao-Ping2,3, XIE Jing1, GUO Qiong-Xia2,   

  1. (1. Fuzhou Airport EntryExit Inspection and Quarantine Bureau, Fuzhou 350209, China; 2. Inspection and Quarantine Center of Fujian EntryExit Inspection and Quarantine Bureau, Fuzhou 350001, China; 3. College of Plant Protection, Fujian Agriculture and Forestry University, Fuzhou 350002, China)
  • Online:2015-04-20 Published:2015-04-20

摘要: 【目的】辣椒实蝇 Bactrocera latifrons (Hendel)为我国重要的检疫性有害生物,其寄主范围广泛,危害严重。由于传统鉴定方法受到饲养周期、饲养条件、虫态等因素的限制,使得果蔬进出口贸易通关速度、疫情快速鉴定受到较大的影响,因此迫切需要开发关于实蝇的快速鉴定识别的技术。【方法】本研究基于mtDNA COI序列设计了一对能够准确鉴定辣椒实蝇的种特异性引物FL680和RL1057,选用辣椒实蝇作为阳性对照,选用番石榴实蝇B. correcta (Bezzi)、桔小实蝇 B. dorsalis (Hendel)和颜带实蝇 B. cilifer (Hendel)等20种实蝇作为阴性对照,进行PCR扩增并将PCR产物进行电泳检测。【结果】仅目标种辣椒实蝇能够扩增出清晰且单一的约378 bp的条带,其余实蝇种类均未出现条带。将本实验建立的种特异性PCR(SS-PCR)鉴定方法应用于实际检疫工作中并得到了验证,表明该方法具有强的种特异性。【结论】本文提出辣椒实蝇快速鉴定识别技术可应用于实蝇的疫情监测和口岸的检疫检测工作。

关键词: 辣椒实蝇, 种特异性引物, 种特异性PCR, mtDNA COI, 快速鉴定

Abstract: 【Aim】 Bactrocera latifrons (Hendel) is one of the most important plant quarantine pests in plant production and trade, and it has wide host range and causes serious damage. Traditional identification methods are limited by such factors as raising period, feeding conditions and developmental stage, thus limiting the clearance rate of customs and quick identification of infestation situation of fruits. For this reason, a rapid identification technique for fruit fly species is desperately needed. 【Methods】 A pair of primers, FL680 and RL1057, was designed and synthesized based on the mtDNA COI sequence. For the PCR amplification and gel electrophoresis, B. latifrons (Hendel) was chosen as the positive control and other 20 species of fruit flies including B. correcta (Bezzi), B. dorsalis (Hendel) and B. cilifer (Hendel) were used as the negative controls. 【Results】 A clear and single 378 bp band was amplified only from B. latifrons, but not from the other 20 fruit flies. The SS-PCR identification method established in this experiment was applied and verified in inspection and quarantine, indicating that this method has high species specificity. 【Conclusion】 The rapid identification method developed here for B. latifrons can be applied in infestation monitoring and quarantine surveillance of fruit flies in ports.

Key words: Bactrocera latifrons, species-specific primers, species-specific PCR (SS-PCR), mtDNA COI, rapid identification