丁型肝炎病毒基因组RNA中核酶区的cDNA的体外转录

In Vitro Transcription of cDNA of Hepatitis Delta Virus Genomic RNA Ribozyme

以质粒ｐＳＶＬＤ３为模板，通过聚合酶链反应（ＰＣＲ）扩增得到１条１３９ｂＰ的ｃＤＮＡ片段，它含有丁型肝炎病毒（ＨＤＶ）基因组ＲＮＡ中核酶（Ｒｉｂｏｚｙｍｅ）区的ｃＤＮＡ，将此片段克隆到ＰＧＥＭ－３２中，经筛选、鉴定，得到克隆株ｐＨＤＶ１０８，提取质粒后经双脱氧末端终止法测序，发现有２个碱基变异。以该质粒为模板，通过Ｔ７ＲＮＡ聚合酶转录出ＨＤＶ基因组ＲＮＡ中核酶区的前体，并观察到其自身裂解产物，自裂率达７１％。

With pSVLD3 as template, a 139hp cDNA fragment containting the region related to hepatitis delta virus(HDV) genomic RNA ribozyme was amplified by polymerase chain reaction(PCR). The PCR products were cloned into pGEM-3Z.A clone(pHDV108) was obtained. The inserted DNA sequencing was carried out by the dideoxy chain termination method. a two-base-mutation was found. With pHDV108 as template, the precursor RNA of HDV genomic RNA ribozyme was obtained by in vitro transcription using T7 RNA polymerase and its self-cleavage rate was 71 %.