应用TaqMan-MGB探针进行松材线虫的实时荧光定量检测技术研究

 松材线虫是国际公认的最重要的检疫性有害生物之一,也是我国2类检疫危险性有害生物,我国口岸多次从货物的木质包装中截获该线虫。由于松材线虫与拟松材线虫在形态上极其相似,难以区分,幼虫更无法用于鉴定。传统的形态学鉴定、生化以及其他分子技术等方法存在费时、准确度不高、灵敏度低等缺点,不易形成标准。我们设计筛选一对引物以及一条MGB探针,对松材线虫进行实时荧光PCR检测。建立了一条从1 pg到10⁴pg标准曲线,相关系数r=0.965。该检测方法省时、准确、快速、无污染。

Quantity assay of Bursaphelenchus xylophilus with TaqMan-MGB probe

Pine wood nematode (Bursaphelenchus xylophilus), one of the most important quarantine pests in the world, is also in the A2 list concerned to China. Entry port of China intercepted it for hundreds times from wood packaging materials of goods. Due to its morphological similarity with Bursaphelenchus mucronatus, it is difficulty to distinguish it from B. mucronatus. Many attempts to develop detecting and diagnosing methods for Bursaphelenchus xylophilus had been conducted, including biochemical technology and molecular technology. However, those were time consuming and instable. A real-time quantitative polymerase chain reaction method using the MGB probe and 1 pair of primers for detecting Bursaphelenchus xylophilus was developed in this study. A liner standard curve was obtained between 1 pg and 104pg pre reaction (r =0.965). The detection was quick, sensitive and little contamination. A faster and practicable method for detecting and diagnosis of B. xylophilus for the quarantine purpose was to be developed.