Nigro Olivia, Alonso Aguirre A, Lu Yuanan
Retrovirology Research Laboratory, Pacific Biomedical Research Center, University of Hawaii at Manoa, Honolulu, HI 96822, USA.
J Virol Methods. 2004 Sep 1;120(1):107-12. doi: 10.1016/j.jviromet.2004.04.011.
Because newly identified green turtle herpesvirus (GTHV) is associated pathogenically with marine turtle fibropapillomatosis (FP) and it has not been isolated in vitro, molecular sequencing and analysis of the genomic DNA of this putative reptilian herpesvirus will enhance the current understanding of GTHV in causing the FP disease. An inverse polymerase chain reaction (IPCR) genomic walking technique was developed to obtain new DNA sequences based on a portion of known genomic sequence. Through two genomic walks, a 2169 bp DNA fragment of GTHV was cloned and sequenced. Sequence analysis shows that this DNA fragment contains the entire gene of the UL28, as well as the partial genomic sequence of the UL27 gene. The UL28 gene is 2250 bp long and encodes a 750-amino acid peptide known as ICP18.5 assembly protein of herpesviruses. Phylogenetic analysis of the GHTV UL28 gene showed a high sequence homology with the UL28 homologs of other herpesviruses and supports the current classification of GTHV to be a member of Alphaherpesvirinae. Identification of the genomic sequences of GTHV provides a molecular base for the development of diagnostic immunoassay and also for the determination of the pathogenic role of GTHV infection.
由于新发现的绿海龟疱疹病毒(GTHV)在致病性上与海龟纤维乳头瘤病(FP)相关,且尚未在体外分离出来,对这种假定的爬行动物疱疹病毒的基因组DNA进行分子测序和分析,将增进目前对GTHV引发FP疾病的了解。开发了一种反向聚合酶链反应(IPCR)基因组步移技术,以基于已知基因组序列的一部分获得新的DNA序列。通过两次基因组步移,克隆并测序了GTHV的一个2169 bp的DNA片段。序列分析表明,该DNA片段包含UL28的完整基因以及UL27基因的部分基因组序列。UL28基因长2250 bp,编码一种750个氨基酸的肽,称为疱疹病毒的ICP18.5装配蛋白。对GHTV UL28基因的系统发育分析表明,它与其他疱疹病毒的UL28同源物具有高度的序列同源性,并支持目前将GTHV分类为α疱疹病毒亚科的成员。GTHV基因组序列的鉴定为诊断免疫测定的开发以及确定GTHV感染的致病作用提供了分子基础。