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目的简化麻疹病毒H1和H2基因型快速鉴定方法的步骤,使之操作更加简单快速。方法将逆转录-套式聚合酶链反应(Reverse Transcription-Nested Polymerase Chain Reaction,RT-nPCR)改进为一步法RT-PCR,使用原方法中的一对内引物,将扩增后的PCR产物用限制性片段长度多态性分析进行基因型鉴定,同时电泳检测也用较安全的荧光染料GeneFinder代替了有致癌性的溴化乙锭染色。结果该研究中使用的已知为H1基因型的3株麻疹病毒,其核酸全部能被改进的一步法RT-PCR有效扩增,并且PCR产物被内切酶SalⅠ有效切开,而且保持了原方法较好的敏感性和特异性。结论改进的麻疹病毒H1和H2基因型快速鉴定方法是一种更加快速、简便又实用的基因定型方法。
Objective To simplify the procedure for rapid identification of the H1 and H2 genotypes of measles virus, making it easier and faster to operate. Methods The reverse transcription-nested polymerase chain reaction (RT-nPCR) was modified to one-step RT-PCR. A pair of primers in the original method was used to amplify the amplified PCR products with Restriction fragment length polymorphism analysis genotype identification, electrophoresis test also used a more secure fluorescent dye GeneFinder instead of carcinogenicity of ethidium bromide staining. Results The three measles viruses known to be H1 genotype used in this study all had their nucleic acids efficiently amplified by the improved one-step RT-PCR and the PCR product was effectively cleaved by the restriction enzyme SalI and maintained the original Method is better sensitivity and specificity. Conclusion The rapid method of rapid identification of the H1 and H2 genotypes of measles virus is a more rapid, simple and practical genotyping method.