中文摘要：

目的：以RNAi技术寻找抑制大鼠Mecp2基因表达的有效序列。方法：首先构建大鼠Mecp2基因与红色荧光蛋白（RFP）的融合基因表达质粒，然后分别与针对大鼠Mecp2基因的4个备选小干扰RNA（siRNA，small interference RNA）共转染HEK293T细胞，构成外源基因系统，以此对于4个备选的siRNA对大鼠Mecp2基因的抑制有效性进行筛选。然后将筛选到的有效序列siRNA转染自身表达大鼠Mecp2基因的PC12细胞，以免疫荧光染色显示大鼠Mecp2蛋白的表达情况，通过内源大鼠Mecp2基因表达被抑制的情况验证所筛选序列的RNA干扰有效性。结果：针对大鼠Mecp2基因mRNA918—936位核苷酸的siRNA（序列：5＇-GCUGUGAAGGAAUCUUCUA-3’）是对大鼠Mecp2基因进行RNA干扰的有效序列。此有效siRNA序列所在的位置相当于大鼠Mecp2基因第4外显子，推测可同时抑制大鼠Mecp2α和大鼠Mecp2β的表达。且此靶序列位于大鼠Mecp2基因编码区，推测可同时抑制1．9kb和10kh的大鼠Mecp2基因mRNA的表达。结论：本研究为建立稳定的大鼠神经元Mecp2基因表达敲低的神经细胞模型和研究脑发育过程中Mecp2基因的功能打下了基础。

英文摘要：

Objective ：To find the valid siRNA （small interference RNA） sequence to knock down rat Mecp2 expression for the analysis of Mecp2 function by RNA interference （RNAi）. Methods： A plasmid （pMecp2-RFP） expressing rat Mecp2 and marker gene red fluorescent protein （RFP） as a fusion gene was constracted. We first selected a candidate valid sequence by cotransfecting the pMecp2-RFP with 4 candidate siRNAs targeting rat Mecp2 to HEK293T cells respectively, and then identified the RNAi efficiency of the candidate valid siRNA by detecting its effect on suppressing the expression of endogenous rat Mecp2 in PC12 cells. Results： siRNA sequence （5＇-GCUGUGAAGGAAUCUUCUA-3＇） targeting rat Mecp2 mRNA 918-936nt was the valid sequence to knock down rat Mecp2 expression by RNA interference. As the target sequence was located in exon 4 of rat Mecp2 mRNA, we assumed that it could suppress the expression of Mecp2α and Mecp2β at the same time. And as the target sequence was located in the coding region of rat Mecp2, we assumed that it could suppress the expression of both 1.9 kb and 10 kb of rat Mecpz transcript. Conclusion：This study has laid the foundation for constructing rat Mecp2 knocked-down neuronal cell model to study the gene function of Mecp2 in brain development.