Fernandez-Cassi Xavier, Rusiñol Marta, Martínez-Puchol Sandra
Department of Genetics, Microbiology and Statistics, Faculty of Biology, University of Barcelona, Barcelona, Spain.
Methods Mol Biol. 2018;1838:173-188. doi: 10.1007/978-1-4939-8682-8_13.
The protocol presented here allows the isolation, purification, nucleic acid extraction, and amplification of DNA/RNA from viruses present in human sera samples. The method allows the random amplification of the viral genomes present by using a Sequence-Independent, Single-Primer Amplification (SISPA) approach enabling the study of both DNA/RNA viruses. An amplification step is needed, as the concentration of viral DNA/RNA in serum samples is low for direct library preparation. The application of the described protocol guarantees enough randomly amplified double-strand DNA for further library preparation using Nextera XT kit from Illumina.
本文介绍的方案可用于从人血清样本中的病毒中分离、纯化、提取核酸以及扩增DNA/RNA。该方法通过使用序列非依赖性单引物扩增(SISPA)方法对存在的病毒基因组进行随机扩增,从而能够研究DNA/RNA病毒。由于血清样本中病毒DNA/RNA的浓度较低,无法直接用于文库制备,因此需要进行扩增步骤。应用所描述的方案可确保获得足够的随机扩增双链DNA,以便使用Illumina公司的Nextera XT试剂盒进行进一步的文库制备。