人源蓝氏贾第虫(Giardia lamblia)病毒部分基因的克隆及序列分析

目的：从我国蓝氏贾第虫（Giardia lamblia)中寻找贾第虫病毒。方法 对人源贾第鞭毛虫北京株进行了体外纯培养，将其总核酸电泳，并用DNA酶和RNA酶处理。根据已发表的蓝氏贾第虫病毒基因序列合成一对引物并进行RT－PCR，将产物回收后连接到Pmd18-T载体上进行克隆并测序，通过BLAST对Gen Bank进行同源性搜索。在贾第虫总核酸电泳图谱上观察到一个分子量约7．0kb的片段。该核酸不能被DNA酶（100μg/ml）降解。但可被RNA酶（10μg/ml）降解。经RT－PCR扩增后得到1条预计856bp的片段，所得序列与蓝氏贾第虫病毒基因同源性为98％。结论 在我国人源贾第鞭毛虫北京株中发现贾第虫病毒，该病毒属于蓝氏贾第虫病毒。

Cloning and sequence analysis of a partial gene of the Giardiavirus

Objective To determine whether there is virus in Giardia lamblia isolated from human in China. Methods Gisrdia lamblia BEIJ88/BTMR1/1 was cultured in vitro. Total nucleic acids from the trophozoites of Giardia lamblia were extracted and used as template for the amplification of the viral genome using RT-PCR method. According to the published nucleotide sequence of Giardiavirus the pair of primers for RT-PCR were designed. The RT-PCR product was cloned pMD18-T vector, and sequenced. Results Apart form the total genomic DNA, a 7.0 kb band was amplified by RT-PCR Electrophoresis of the total nucleic acid extracted from the Giardia lamblia isolate revealed that apart from the large genomic DNA, there was a 7.0kb nucleic acid band which was sensitive to RNase A digestion. A fragment of 856bp was amplified by RT-PCR and cloned. The DNA sequence of the cloned fragment was nearly identical with the published sequence of the Giardia lamblia BEIJ88/BTMR1/1 virus. Conclusion The dsRNA virus in Giardia lamblia BEIJ88/BTMR1/1 was detected in the Giardia lamblia isolate from human in China.