菜豆荚斑驳病毒免疫捕获一步RT-PCR检测

Detection of Bean Pod Mottle Virus by One-step IC-RT-PCR

【研究目的】建立快速、准确、简便的检测大豆种子上菜豆荚斑驳病毒的免疫捕获一步RT-PCR方法(一步IC-RT-PCR)。【方法】采用抗原捕获和一步RT-PCR相结合的方法,并通过反应条件优化,确定一步IC-RT-PCR的反应程序,同时对该方法的特异性及实际检测效果进行验证。【结果】成功建立了一步IC-RT-PCR检测菜豆荚斑驳病毒方法,能够从感染菜豆荚斑驳病毒的大豆种子上扩增出预期大小的特异性片段,而对其他病毒及健康大豆无特异性反应;应用该方法对不同产地大豆种子样品进行检测,结果从来自美国的大豆样品中检出菜豆荚斑驳病毒,检出率为50%。【结论】一步IC-RT-PCR方法可以快速、准确地检测大豆种子上的菜豆荚斑驳病毒,适用于口岸检验检疫。

[OBJECTIVE]Establishment of a rapid, accurate and sensitive one-step IC-RT-PCR method to detect Bean pod mottle virus （BPMV） in soybean seed. [METHOD]One-step IC-RT-PCR was developed to detect BPMV by using RT-PCR in combination with immunocapture. After optimization of the reaction conditions, One-step IC-RT-PCR procedure was established, and then the specificity and practical effect of detection were validated. [RESULTS]The One-step IC-RT-PCR could successfully amplify target fragment from BPMV-infected soybean seeds while not from other virus infected and healthy soybean seeds. BPMV in soybean from different producing areas was detected by means of the One-step IC-RT-PCR. The results showed that BPMV was detected in soybean seed from America and the incidence was 50%.[CONCLUSION]The One-step IC-RT-PCR was rapid and accurate for detection of BPMV in soybean seed, which can be applied to the field of port inspection and quarantine.