烟草赤星病菌实时荧光定量PCR检测方法的建立

Development of real-time fluorescence quantification PCR system for detection of Alternaria alternata in tobacco leaves

摘要: 为实现对烟草叶片中烟草赤星病菌的SYBR Green I实时定量PCR（q-PCR）快速检测，根据链格孢属ITS基因序列差异位点，设计了1对特异性引物。通过构建含有目的片段的重组质粒，建立了标准曲线，构建了q-PCR快速定量检测方法并对感病叶片进行检测，灵敏度达到1.0×10^-3 ng/μL。实验样品验证表明, 该方法可以对烟草赤星病菌进行快速检测和定量。

Abstract: In order to developed a SYBR Green I real-time fluorescence quantitative PCR system for rapidly detecting Alternaria alternata in tobacco leaves, a pair of specific primers were designed according to the sequence differences of ITS of Alternaria.By constructing a recombinant plasmid containing the targeted fragments, the q-PCR rapid detection system was developed with its standard curve to detect A. alternata-infected tobacco leaves. The sensitivity of detection was above 1×10^-3 ng/μL, which indicated that this system could ensure efficient detection and quantification of A. alternata in tobacco leaves.