核酶抑制N_2a细胞血栓素合酶mRNA的表达

Inhibition of Hammerhead Ribozyme against N_2a TXS mRNA Expression

根据“锤头状结构”核酶的设计原理 ,以小鼠全长血栓素合酶 (TXS) m RNA为靶系列 ,计算机预测其二级结构 ,设计核酶 ,并人工合成核酶对应的 DNA片段 ,克隆入真核表达载体 p EGFP- C1,经脂质体转染 N2 a细胞 ,用原位杂交的方法检测 TXS m RNA的表达水平。结果表明 :针对小鼠 TXS m RNA的核酶 R15、R5 44 ,对内毒素刺激的 N2 a细胞 TXS m RNA的表达具有显著的抑制作用 ,核酶 R15、R5 44可望用于基因治疗 ,抑制病理条件下 TXS m

The possible secondary structures of RNA sequence of TXS strain which was taken as the targeting RNA was predicted, the ideal cleavage sites was selected and the ribozymes were designed by computer according to the principle of 'hammerhead structure' and lowest energy. Then, hammerhead ribozyme DNA was cloned into pEGFP-C1 and delivered into N 2a cell by liposomes. The expression of TXS mRNA was detected by using in situ hybridization. The results showed ribozymes R15 and R544 against murine TXS mRNA could markedly inhibit the expression N 2a TXS mRNA stimulated by endotoxin. It is hopeful that R15 and R544 might be a good inhibitor of TXS in gene therapy.