Bi S L, Purdy M A, McCaustland K A, Margolis H S, Bradley D W
Hepatitis Branch, Centers for Disease Control and Prevention, Atlanta, GA 30333.
Virus Res. 1993 Jun;28(3):233-47. doi: 10.1016/0168-1702(93)90024-h.
In this study an IgM antibody-mediated antigen-capture procedure for direct extraction of hepatitis E virus (HEV) RNA from clinical specimens was developed and used with an efficient method for generating viral cDNA that was subsequently sequenced using the dideoxy chain termination method. This is the first time the complete HEV genome has been isolated directly from a single human clinical specimen obtained during an outbreak of enterically transmitted non-A, non-B hepatitis. When the Chinese-derived sequence was compared with the original isolate of Burmese HEV from an experimentally infected cynomolgus macaque, the homology between the two sequences was 94% and 98.5% at the nucleotide and amino acid levels, respectively. The methods we developed for generating and sequencing genomic HEV cDNA dramatically improved the efficiency of cloning the viral genome and should be helpful for continued analysis of this virus as well as other RNA viruses that have proven to be difficult to clone and sequence directly.
在本研究中，开发了一种IgM抗体介导的抗原捕获程序，用于从临床标本中直接提取戊型肝炎病毒（HEV）RNA，并与一种高效生成病毒cDNA的方法一起使用，随后使用双脱氧链终止法对其进行测序。这是首次直接从肠道传播的非甲非乙型肝炎暴发期间获得的单个人类临床标本中分离出完整的HEV基因组。当将源自中国的序列与来自实验感染食蟹猴的缅甸HEV原始分离株进行比较时，两条序列在核苷酸和氨基酸水平上的同源性分别为94%和98.5%。我们开发的用于生成和测序基因组HEV cDNA的方法显著提高了克隆病毒基因组的效率，并且应该有助于对该病毒以及其他已证明难以直接克隆和测序的RNA病毒进行持续分析。
