传染性支气管炎病毒的部分纯化及随后用于下一代测序的病毒RNA分离。
Partial purification of IBV and subsequent isolation of viral RNA for next-generation sequencing.
作者信息
Keep Sarah M, Bickerton Erica, Britton Paul
机构信息
The Pirbright Institute, High Street, Compton, Newbury, Berkshire, RG20 7NN, UK,
出版信息
Methods Mol Biol. 2015;1282:109-12. doi: 10.1007/978-1-4939-2438-7_11.
RNA viruses are known for a high mutation rate and rapid genomic evolution. As such an RNA virus population does not consist of a single genotype but is rather a collection of individual viruses with closely related genotypes-a quasispecies, which can be analyzed by next-generation sequencing (NGS). This diversity of genotypes provides a mechanism in which a virus population can evolve and adapt to a changing environment. Sample preparation is vital for successful sequencing. The following protocol describes the process of generating a high-quality RNA preparation from IBV grown in embryonated eggs and then partially purified and concentrated through a 30% sucrose cushion for NGS.
RNA病毒以高突变率和快速的基因组进化而闻名。因此，RNA病毒群体并非由单一基因型组成，而是由具有密切相关基因型的单个病毒组成的集合——即准种，可通过下一代测序（NGS）进行分析。这种基因型的多样性提供了一种机制，病毒群体可借此进化并适应不断变化的环境。样本制备对于成功测序至关重要。以下方案描述了从在鸡胚中生长的传染性支气管炎病毒（IBV）中制备高质量RNA的过程，然后通过30%蔗糖垫层进行部分纯化和浓缩，以用于NGS。
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