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目的 研究锤头状核酶对雄激素受体 (AR)mRNA片段的体外切割活性 ,探讨抗雄激素基因治疗的新途径。方法 计算机模拟ARmRNA片段二级结构 ,选择第 10 18位GUC为核酶切割位点 ,设计特异性切割AR的锤头状核酶 (RZ) ,合成含T7启动子的核酶和底物模板链 ,体外转录RNA ,37℃条件下行核酶体外切割反应。结果 核酶在体外 37℃反应 1h ,可将 98%的底物RNA ( 2 5 5nt)切割为 16 8nt和 87nt2个片段。结论 特异性切割AR的锤头状核酶在体外对底物RNA有良好的切割活性 ,为应用核酶阻断雄激素受体基因表达奠定了基础。
Objective To study the in vitro cleavage activity of androgen receptor (AR) mRNA by hammerhead ribozyme and to explore a new way of anti-androgen gene therapy. Methods The secondary structure of ARmRNA fragment was simulated by computer. The site of GUC cleavage at position 10 18 was selected. The specific ribozyme (RZ) was designed to cut AR, and the ribozyme containing T7 promoter and substrate template chain , In vitro transcription of RNA, 37 ℃ under the conditions of the ribozyme in vitro cleavage reaction. As a result, the ribozyme was reacted at 37 ° C for 1 h in vitro, and 98% of substrate RNA (25 5 nt) was cut into 16 8 nt and 87 nt 2 fragments. Conclusion The hammerhead ribozyme specific for AR cutting has good cleavage activity on substrate RNA in vitro, which lays the foundation for the use of ribozyme to block androgen receptor gene expression.