摘要
自上海市及江苏省慢性丙型肝炎患者血清中分离丙型肝炎病毒(HCV)RNA,采用自行设计的引物,经逆转录-聚合酶链反应(RT-PCR),获得长为694bp的C33c抗原基因cDNA,将其克隆于表达载体并作序列分析。结果表明,上海株HCV-S1及江苏株HCV-S2该区域核苷酸水平有很高的同源性(>99%),上海及江苏株HCV与美国原型HCV-1、日本株HCV-J及河北株HCV-HB在该区域核苷酸水平的差异分别为18.64%、5.55%及8.01%~8.17%,氨基酸水平的差异在2.31%~6.02%之间。上海株同一份血清三个独立克隆间核苷酸水平有0.6%~0.9%的差异。本实验为进一步表达C33c蛋白作好了准备。
Sera from two chronic hepatitis C patients were used for isolation of HCV RNA, one from Shanghai and the other from Jiangsu Province.Using primers designed by ourself,a 694bp cDNA fragment of C33c protein gene located in NS3 region of HCV was generated by RT-PCR. After cloning into the expressing vector pBluescript Ⅱ, the fragments were sequenced by the dideoxynucleotide method.The cDNA fragment of Shanghai isolate HCV-S1 and Jiangsu isolate HCV-S2 showed a high identity(>99%) at the nucleotide(nt) level.When the two isolates were compared with the original isolate of US HCV-1、Japanese isolate HCV-J and Hebei isolate HCV-HB in the examined region,18.64%、5.55% and 8.01~8.17% diversity at the nucleotide level was found respectively and 2.31~6.02% diversity was found at the predicted amino acid (aa) level. In the study,0.6%~0.9% diversity at the nucleotide level was observed among three independent clones obtained from a single serum sample.The study here also served as a preparation for further expression of C33c protein.
出处
《上海第二医科大学学报》
CSCD
1996年第2期148-152,共5页
Acta Universitatis Medicinalis Secondae Shanghai
关键词
丙型肝炎病毒
CDNA克隆
序列分析
hepatitis C virus, polymerase chain reaction, cDNA cloning, sequence analysis