中文摘要：

采用自行设计5’固定3’随机的文库对基因mRNA进行杂交用于逆转录反应以筛选mRNA的寡核苷酸结合靶点。对人I型跨膜糖蛋白-血型糖蛋白A（glycophorinA，GPA）的mRNA筛选了4个反义寡核苷酸可结合靶点，分别设计反义核酸（Antisense），分别加入mRNA中用RNaseH验证各靶点的核酸结合和切割效率，最终确定2个高效结合和切割靶点。再设计Ribozyme，构建表达核酶（Ribozyme）质粒，利用慢病毒（Lentivirus）包装技术，感染人源红系白血病细胞株K562细胞，在细胞水平验证其下调GPA基因表达的效果，对转染细胞mRNA进行反转录和RealTimePCR分析mRNA表达水平，并在蛋白水平进行了WesternBlot分析。结果表明文库结合逆转录方法筛选靶点设计的Ribozyme具有高效率下调膜受体表达的作用，GPA为I型跨膜糖蛋白，该实验为筛选mRNA靶点提供参考方法，并对膜受体表达干预有参考价值。

英文摘要：

An oligodeoxynucleotide library which the sequence were defined at 5 prime and randomized at 3 prime was employed to screen mRNA accessible targets, in reverse transcription and PCR after hybridized the library with mRNA. The mRNA of Glycophorin A （ GPA）, type I transmembrane glyeoprotein, was screened and obtained 4 targets sequences. Accordingly 4 antisense nucleic acids designed respectively, the binding efficiency of every target were verified by using RNase H with antisense nucleic acids. Among them 2 targets showed better effects on binding and cutting. Designed 2 ribozymes to these targets, packaged in lentivirus system, then infected K562 cells（ human erythroid leukemia line） , the down-regulation effect of gene expression was validated by Real Time RT-PCR and by Western Blot. It was found that the screened targets showed the best effective knocking down effects on gene expression. The study provided a reference for mRNA targets screening and Ribozyme design, and was helpful in membrane receptor expression interference, since GPA is a transmembrane protein.