水中诺如病毒富集效果及核酸提取方法的比较

目的比较诺如病毒检测时3种核酸提取方法的效率,并对水中诺如病毒的检测方法进行初步研究。方法采用Trizol法、硅胶柱法及磁珠法分别提取含有诺如病毒的粪便样本的核酸,Real-time PCR检测其Ct值,比较3种方法提取核酸的效率。将含有诺如病毒的粪便加入纯水中,采用MCE-PEG法富集后,Real-time PCR检测其Ct值,评价MCE-PEG法富集病毒的效果。结果硅胶柱法的Ct平均值最小,在3种方法中提取诺如病毒RNA效率最高,磁珠法提取效果次之,Trizol法效果最差;MCE-PEG法富集病毒效果良好。结论磁珠法提取诺如病毒RNA效率虽不是最高,但可实现自动化,节省工作时间,降低了人力成本,是3种方法中基层实验室提取诺如病毒RNA最适合的一种;MCE-PEG法可以用来富集水中的诺如病毒。

Concentration of Noroviruses from Water and Extraction of Nucleic Acids

Objective To compare the efficacies of three nucleic acid extraction methods for the detection of noroviruses,and to preliminarily investigate the methods for the detection of noroviruses in water. Methods Nucleic acids were extracted from stool samples containing noroviruses using Trizol reagent,silica gel column and magnetic beads,respectively. The efficacies of the three methods for nucleic acid extraction were compared. Stool samples containing noroviruses were added to pure water. After enrichment using MCE-PEG method,the cycle threshold（ Ct） values were obtained by real-time PCR to evaluate the efficacy of MCE-PEG method for enrichment of viruses. Results The minimum average Ct value was achieved by silica gel column method,which was the most effective for the extraction of norovirus nucleic acids. The silica gel column method and magnetic bead method exhibited the moderate and poor efficacy for the extraction of norovirus nucleic acids,respectively. The MCE-PEG method was effective for the enrichment of viruses. Conclusion Although magnetic bead method is not the most effective for the extraction of norovirus nucleic acids,it can realize automation,save time and reduce labor costs.Therefore,magnetic bead method is the most suitable for norovirus nucleic acid extraction in grassroots laboratory. The MCE-PEG method can be used to enrich noroviruses in water.