Li Zhuo, Yu Meng, Zhang Hong, Wang Hai-Yan, Wang Lin-Fa
Renal Division and Institute of Nephrology, Peking University First Hospital, Beijing, China.
J Virol Methods. 2005 Dec;130(1-2):154-6. doi: 10.1016/j.jviromet.2005.06.022. Epub 2005 Aug 1.
Rapid amplification of cDNA ends (RACE) is a powerful PCR-based technique for determination of RNA terminal sequences. However, most of the RACE methods reported in the literature are developed specifically for the mapping of eukaryotic transcripts with 3' poly-A tail and 5' cap structure. In this study, an improved RACE strategy was developed which allows both 5' and 3' RACE of paramyxovirus genomic RNA using the same set of common molecular biology reagents without having to rely on expensive RACE kits. Mapping of RNA genome terminal sequences is an essential part of characterizing novel paramyxoviruses since these sequences contain important signals for genome replication and transcription, and are important molecular markers for studying virus evolution. The usefulness of this strategy was demonstrated by rapid characterization of both genome ends for a novel paramyxovirus recently isolated from human kidney primary cells. The RACE strategy described in this paper is simple, cost-effective and can be used to map genome ends of any RNA viruses.
cDNA末端快速扩增（RACE）是一种基于聚合酶链式反应（PCR）的强大技术，用于确定RNA的末端序列。然而，文献中报道的大多数RACE方法都是专门为绘制具有3' 聚腺苷酸尾和5' 帽结构的真核转录本而开发的。在本研究中，开发了一种改进的RACE策略，该策略允许使用同一套常见的分子生物学试剂对副粘病毒基因组RNA进行5' 和3' RACE，而无需依赖昂贵的RACE试剂盒。绘制RNA基因组末端序列是鉴定新型副粘病毒的重要组成部分，因为这些序列包含基因组复制和转录的重要信号，并且是研究病毒进化的重要分子标记。最近从人肾原代细胞中分离出的一种新型副粘病毒的基因组两端的快速鉴定证明了该策略的有效性。本文所述的RACE策略简单、经济高效，可用于绘制任何RNA病毒的基因组末端。
