锤头状核酶阻断雄激素受体基因表达的研究

The hammerhead ribozyme-induced blocking effects on gene expression of androgen receptor

目的 探讨核酶 (RZ)在细胞水平调节雄激素受体 (AR)基因表达的作用。方法 设计并合成针对AR的锤头状核酶序列 ,分子克隆技术构建活性核酶和无活性核酶表达载体 ,脂质体介导下转染雄激素受体表达阳性细胞株T47D。转染第 1～ 3天 ,应用免疫组织化学和逆转录 聚合酶链反应 (RT PCR)技术检测细胞内AR基因表达水平。结果 AR活性核酶表达载体转染后 ,T47D细胞ARmRNA水平降低 2 2 .4%～ 50 .7% (P <0 .0 5) ,AR蛋白阳性细胞减少 7.1 0 %～1 1 .90 % (P <0 .0 5)。而无活性核酶对细胞AR基因表达差异无显著性 (P >0 .0 5)。结论 成功构建AR锤头状核酶表达载体 ,能特异性切割ARmRNA 。

Objective To explore the regulatory effects of ribozyme on the cellular gene expression of androgen receptor (AR). Methods The specific hammerhead ribozyme sequences for AR gene were designed and synthesized. The active and inactive ribozyme expression vectors were constructed by molecular cloning techniques and transferred into AR positive cell line T47D with liposome. After being transferred for 1 3 days, the cellular AR gene expression was detected by RT PCR and immunohistochemical methods. Results When T47D cells were transferred by AR specific ribozyme, the AR mRNA level was reduced by 22.4%-50.7% (P<0.05), with the AR protein positive cells being decreased by 7.10%-11.90% (P<0.05). However, there was no obvious effects on cellular gene expression of AR by inactive ribozyme (P>0.05).Conclusion The hammerhead ribozyme expression vector was successfully constructed, which could block the gene expression of AR by specific cutting effects on AR mRNA.