【摘要】 草莓白化伴随病毒（strawberry pallidosis-associated virus, SPaV）能引起草莓白化病，与其他病毒复合侵染后会加重症状。本研究建立了SPaV的反转录环介导等温扩增方法（reverse transcription-loop-mediated isothermal amplification, RT-LAMP）,对反应体系的各条件进行评估优化后，检测了RT-LAMP的特异性和灵敏度，且对田间样品进行了测试。结果表明，优化后的反应体系包含2.5μL 10×Isothermal Amplification Buffer、6 mmol/L Mg²⁺、1 mmol/L dNTPs、1μmol/L SPaV-FIP和SPaV-BIP,0.1μmol/L SPaV-F3和SPaV-B3,0.3μmol/L SPaV-LB,0.4 mol/L Betaine, 0.256 U/μL WarmStart Bst 2.0 DNA Polymerase, 0.12 U/μL WarmStart RTx Reverse Transcriptase, 1μL RNA,反应条件为61℃恒温孵育40 min。采用优化后的反应体系，在特异性检测中，只有携带了病毒SPaV的样品才能发生阳性扩增。灵敏度测试中，RT-LAMP比RT-PCR灵敏度高10倍。田间应用中，RT-LAMP和RT-PCR检测结果一致。本研究建立的SPaV RT-LAMP检测方法操作简便、快速、特异，可供实验室及生产实践中应用，助力脱毒种苗的鉴定和田间病毒病调查。更多还原

【Abstract】 Strawberry pallidosis-associated virus（SPaV） can cause strawberry pallidosis disease, and the symptoms will worse when SPaV coinfects with other viruses. This study established a reverse transcription-loop-mediated isothermal amplification（RT-LAMP） for SPaV detection. After evaluating and optimizing the conditions of the reaction system, the specificity and sensitivity of this RT-LAMP procedure were mensurated and field samples were tested. The results showed that the optimized reaction system contained 2.5 μL 10×Isothermal Amplification Buffer, 6 mmol/L Mg²⁺, 1 mmol/L dNTPs, 1 μmol/L SPaV-FIP and SPaV-BIP, 0.1 μmol/L SPaV-F3 and SPaV-B3, 0.3 μmol/L SPaV-LB, 0.4 mol/L Betaine, 0.256 U/μL WarmStart Bst 2.0 DNA Polymerase, 0.12 U/μL WarmStart RTx Reverse Transcriptase, 1 μL RNA,and incubated at 61℃ for 40 min. Only the sample containing the SPaV could be positively amplified by the optimized reaction system. The sensitivity of this RT-LAMP was 10 times higher than that of RT-PCR detection. In the field application, the RT-LAMP detection result was consistent with RT-PCR. The RT-LAMP method developed for SPaV detection in this study is simple, rapid and specific, and can be used in laboratory and field to identify virus-free seedlings and investigate virus diseases.更多还原