中文摘要：

目的构建小鼠巨噬细胞半乳糖特异性凝集素（GSL）靶向的RNAi质粒载体,研究其对小鼠和巨噬细胞GSL基因表达的沉默作用。方法根据GSL核苷酸序列,合成3条特异性双链小干扰RNA（siRNA）分子,退火形成双链,分别连接到RNAi-Readyp SIREN-RetroQ-ZsGreen Vector,转化大肠杆菌得到阳性克隆,经测序鉴定确认siRNA载体构建成功后,转染小鼠巨噬细胞RAW264.7及尾静脉注射BALB/c小鼠,应用实时定量PCR和免疫组化法评价siRNA载体对小鼠和巨噬细胞GSL基因表达的抑制作用。结果 siRNA载体对小鼠体内GSL基因的抑制率为84-96%,对巨噬细胞GSL基因的抑制率可达61.98%～83.02%。结论构建的siRNA载体能有效抑制目的基因在体内外的表达,该结果为分析半乳糖特异性凝集素在布鲁氏菌侵染过程中的功能奠定了基础。

英文摘要：

To construct plasmid vector of RNAi for galactose-specific lectin （GSL） and observe the silencing effect on gene expression of GSL in mouse and macrophage respectively,three interference sequences were synthesized,annealed and cloned into RNAi-Ready pSIREN-RetroQ-ZsGreen vector. The recombinant plasmids were transformed into E.coli and identified by sequencing. The siRNAs were transfected into RAW264.7 macrophages and injected into BALB/c mice through vena caudalis. The inhibition effects were evaluated in RAW264.7 cells and experimental mice by real-time PCR and immunohistochemistry respectively. The results showed that the GSL genes of mice could be inhibited by siRNA vectors with an inhibition rate of 84%-96%,while the ratio of GSL genes in macrophage inhibited by siRNA vectors was 61.98%-83.02%. The results conveyed that the constructed siRNA vector could strongly inhibit GSL gene expression in vitro and in vivo,which would be helpful to further study on the function of GSL during Brucella invading macrophage.