CAJ | 학술논문

目的研究广州地区先天性感染的人巨细胞病毒(HCMV)临床低传代分离病毒株UL144基因序列的多态性,探讨UL144基因在HCMV致病中的作用.方法对3株经多重PCR鉴定HCMV DNA为阳性的临床低传代分离株进行HCMV UL144基因全序列PCR扩增,PCR产物克隆到pMD18-T载体上再测序,将其序列与GenBank中公布的其它10株临床分离株UL144基因一起进行分析.结果本实验克隆并测序了HCMV临床低传代D3、D2和D52病毒株的UL144基因,提交GenBank,已被GenBank收录,序列号分别为DQ180368、DQ180382和DQ180355.HCMV临床低传代D3、D2和D52病毒株的UL144基因均全长531 bp.通过blast分析,从GenBank中找到了10株HCMV病毒株的UL144与D3、D2和D52的UL144基因具有较高的同源性,经过序列的比对,发现UL144基因DNA序列比较保守,只在4处有变异,且变异均为碱基替换,无插入或缺失,编码蛋白由176个氨基酸残基组成,氨基酸序列也比较保守,各分离株中变异率为1.1%;HCMV UL144编码蛋白翻译后修饰位点在所有分离株中均高度保守;所有分离株UL144蛋白的等电点均为8.97.结论广州地区临床低传代分离株HCMV UL144基因DNA及其编码产物的氨基酸序列是比较保守的,但仍存在一定的多态性.提示UL144基因在先天性感染中可能具有重要作用.
목적 연구엄주지구선천성감염적인거세포병독(HCMV)림상저전대분리병독주UL144기인서렬적다태성,탐토UL144기인재HCMV치병중적작용.방법 대3주경다중PCR감정HCMV DNA위양성적림상저전대분리주진행HCMV UL144기인전서렬PCR확증,PCR산물극륭도pMD18-T재체상재측서,장기서렬여GenBank중공포적기타10주림상분리주UL144기인일기진행분석.결과 본실험극륭병측서료HCMV림상저전대D3、D2화D52병독주적UL144기인,제교GenBank,이피GenBank수록,서렬호분별위DQ180368、DQ180382화DQ180355.HCMV림상저전대D3、D2화D52병독주적UL144기인균전장531 bp.통과blast분석,종GenBank중조도료10주HCMV병독주적UL144여D3、D2화D52적UL144기인구유교고적동원성,경과서렬적비대,발현UL144기인DNA서렬비교보수,지재4처유변이,차변이균위감기체환,무삽입혹결실,편마단백유176개안기산잔기조성,안기산서렬야비교보수,각분리주중변이솔위1.1%;HCMV UL144편마단백번역후수식위점재소유분리주중균고도보수;소유분리주UL144단백적등전점균위8.97.결론 엄주지구림상저전대분리주HCMV UL144기인DNA급기편마산물적안기산서렬시비교보수적,단잉존재일정적다태성.제시UL144기인재선천성감염중가능구유중요작용.
Objective To investigate the polymorphism of human cytomegalovirus (HCMV) UL144 gene of the low passage clinical isolates in Guangzhou and explore the role of UL144 gene in HCMV pathogenicity. Methods The clinical isolates of HCMV were obtained from the urine sample collected from those infants with intra-uterus HCMV infection in Guangzhou. The virus genome DNA was extracted. According to the genome sequence of Toledo, primers for UL144 gene were designed and used to amplify the complete open reading frames (ORF) of the UL144 gene in our 3 different clinical isolates. These ORFs of the UL144 gene were cloned into pMD18-T vector and their sequences were confirmed by sequencing. Bioinformatics methods were used subsequently to analyze the polymorphisms of these genes in different stains. Results Three HCMV low passage clinical isolates were successfully isolated, named D2, D3 and D52. As shown by PCR, all of these three strains contained UL144 ORF region. Three complete ORFs were amplified in total and their sequences were submitted to GenBank (Accession No.: DQ180368, DQ180382 and DQ180355). In D2, D3 and D52 isolates, their UL144 ORFs consisted of 531 nucleotides. DNA sequences were quite conservative,all variability were base substitution, and the amino acid sequences were high conservative, the rate of amino acid variability was 1.1%. There were no additional or deleted sites of posttranslational modification of UL144 protein in all clinical isolates. There were some differences in the secondary structure among different isolates. The isoelectric point of UL144 protein of all clinical isolates was 8.97. Conclusions All DNA and deduced amino acid sequences of UL144 gene share great similarity among Guangzhou HCMV clinical strains regardless of their polymorphism. It implies that maybe UL144 gene plays an important role in congenital infection.