葡萄根癌病菌的快速检测技术

Rapid Molecular Detection of Agrobacterium vitis

根据葡萄根癌病菌核糖体基因间隔片段(16S-23S Ribosomal DNA Intergenic Spacer,ITS)特异区域设计,合成了一对引物,建立了快速检测葡萄根癌病菌的PCR方法,并对反应条件进行优化,组装成快速检测试剂盒。结果表明:只有葡萄根癌病菌能扩增出大小为758 bp的特异性条带,而非葡萄根癌病菌均未能扩增出任何条带。检测灵敏度达10个细菌/μL左右,且稳定性好,经多次反复冻融,扩增条带未发现有明显变化。通过对田间病样进行检测,证实该试剂盒具有操作简便、快速(4 h左右完成检测)、灵敏度高、特异性强、重复性和稳定性好等优点。

To establish a simple,sensitive,accurate and rapid PCR method for detection of Agrobacterium vitis in grape,one pair of specific primers were designed and synthesized according to the conserved 16S-23S ITS available in grape,and reaction parameters were optimized to develop a PCR selection kit.The results showed that the ITS of 758 bp was amplified in all detected grape by the PCR and no any fragments in non-grape species.The sensitivity of PCR was 10 CFU and stability was good.The amplified sequence had nothing abnormity after repeated freezing and thawing.The developed kit was rapid,simple,sensitive,spedific,accurate and easy for detection by field sample testing.