核酶及突变核酶在体外对HBV mRNA的切割作用

Effect of specific ribozyme and mute ribozyme cleave on HBV mRNA in vitro

目的 :探讨多位点核酶对乙型肝炎病毒 (HBV)信使核糖核酸 (mRNA)的切割作用 ,以及保守碱基突变后对切割活性的影响。 方法 :利用计算机辅助设计针对HBVC区基因的多位点核酶及保守碱基突变的双位点核酶 ,构建多位点核酶及双位点突变核酶的自剪切载体 (pGEMRz12 3、pGEMmRz12 ) ,观察核酶及突变核酶对靶RNA的切割作用。 结果 :构建后的多位点串联核酶及突变核酶的自剪切转录载体在体外转录后 ,其顺式核酶可发生自剪切 ,并可将目的核酶释放出来。多位点核酶对HBVmRNA有明确的切割作用 ,而保守碱基突变后对HBVmR NA则无切割作用。 结论 :抗HBV多位点核酶在体外可明确切割HBV靶RNA ,突变后的核酶无切割活性 。

Objectives:To study the cleavage activity of multi target ribozyme anti HBV mRNA, and the effect of conserved base which was mutated to the activity of cleavage. Methods:Three connected ribozyme and mutated ribozyme against hepatitis B virus core gene were designed with computer, then constructed transcripitional vector of multi target ribozyme and mutated ribozyme which containing 5′,3′ self splicing ribozymes. The cleavage activity of ribozyme and mutated ribozyme on target RNA were analysed. Results:The ribozyme can be released from the self splicing vector after transcripitated.The ribozyme can specifically cleave different cleavage sites on the target mRNA.But the muta ribozyme can not cleave the target HBV mRNA. Conclusions:The connected ribozymes with computer design can cleave RNA of hepatitis B virus core gene.It is necessary of the conserved base for the cleavage activity of ribozyme.