摘要
拟松材线虫与松材线虫的亲缘关系极为接近,是松材线虫病病原鉴定的主要干扰物种。为了排除松材线虫病病原鉴定过程中拟松材线虫的干扰,采用TaqMan实时荧光PCR技术,以核糖体DNA内转录间隔区(ITS)为模板设计拟松材线虫特异性引物和TaqMan探针,开展拟松材线虫分子检测和鉴定技术研究。结果表明:拟松材线虫ITS2区域存在较多的碱基差异,以此为模板设计的拟松材线虫引物和TaqMan探针组合具有较强的特异性;该组合灵敏性高,能够实现对拟松材线虫单条线虫甚至单个卵的分子检测和鉴定。结合现有的松材线虫分子鉴定技术,可以为进一步构建松材线虫双重检测技术体系奠定基础,为林业和海关检疫提供可靠的技术支撑。
Bursaphelenchus mucronatus is the major interfering species in the identification of Bursaphelenchus xylophilus since their highly-similar genetic relationship as two sibling species.In order to eliminate the interference of B.mucronatus in the identification of B.xylophilus,the specific primers and TaqMan probes of B.mucronatus were designed by TaqMan real-time PCR,using the ribosomal internal transcribed spacer(ITS)as the template,and the molecular detection and identification of B.mucronatus were studied.There were many base differences in ITS2 region of B.mucronatus,and the primers and TaqMan probe combination designed based on this template had strong specificity.The selected primer pair and probe were reproducible and feasible for detection and accurate identification of a single nematode even single egg of B.mucronatus.Combined with the existing molecular identification techniques of B.xylophilus,the results could lay a foundation for the further construction of the double-detection system of pine wood nematodes,and provide reliable technical support for forestry and customs quarantine.
作者
宋志强
张旭君
康李鹏
王立超
周立峰
陈凤毛
SONG Zhiqiang(Forestry Administration of Chun'an County,Chun'an 311799,China)
出处
《中国森林病虫》
2021年第5期27-31,共5页
Forest Pest and Disease
基金
中国博士后科学基金项目“MicroRNA-239抑制松材线虫胚胎发育研究”(2020M671796)
。
