Nerva Luca, Varese Giovanna C, Turina Massimo
Istituto per la Protezione Sostenibile delle Piante, CNR, Torino, Italy.
Department of Life Sciences and Systems Biology, Mycotheca Universitatis Taurinensis (MUT), University of Turin, Torino, Italy.
Methods Mol Biol. 2018;1746:97-114. doi: 10.1007/978-1-4939-7683-6_8.
Here we describe the protocols to characterize the virome associated to fungi isolated from marine organisms assessed on the seagrass Posidonia oceanica and on the marine animal Holothuria poli. We provide detailed protocols for fungal isolation, fungal growth, and total RNA extraction. Ribosomal RNA depletion, cDNA library synthesis and normalization, and sequencing runs on different platforms are part of the protocols that are generally outsourced and therefore are not described in this chapter. We describe, instead, how raw reads are assembled into contigs and how to search for putative viral sequences. Furthermore, we detail qualitative checks to infer the existence of the virus as a replicative biological entity.
在此，我们描述了用于鉴定与从海洋生物中分离出的真菌相关的病毒群落的实验方案，这些海洋生物是在海草波喜荡草和海洋动物多疣海参上进行评估的。我们提供了真菌分离、真菌培养和总RNA提取的详细实验方案。核糖体RNA去除、cDNA文库合成与标准化以及在不同平台上的测序运行是通常外包的实验方案的一部分，因此本章不再描述。相反，我们描述了如何将原始读数组装成重叠群以及如何搜索假定的病毒序列。此外，我们详细说明了定性检查，以推断病毒作为复制性生物实体的存在。
