日本血吸虫mir-125b靶向调控宿主CD276的鉴定研究
发布时间：2019-03-19 11:17
【摘要】：通过荧光素酶实验验证宿主CD276为日本血吸虫mir-125b的靶基因,并分析感染血吸虫后宿主CD276基因在不同组织中的表达情况。利用PCR扩增mir-125b与CD276互作的核酸序列,并将目的片段克隆至表达荧光素酶报告基因质粒的下游。将重组质粒转染到HEK293T细胞内,检测荧光素酶报告基因表达。进一步利用荧光定量PCR技术检测小鼠巨噬细胞在转染血吸虫mir-125b mimic后,靶基因CD276的表达情况,同时检测感染血吸虫不同时间段、小鼠不同组织中CD276的表达情况。荧光素酶实验表明转染mir-125b mimic和重组质粒后细胞的荧光素酶报告基因的表达极显著降低(P0.01),提示mir-125b可与CD276相关区域互作,抑制报告基因的表达;血吸虫mir-125b mimic转染小鼠巨噬细胞后导致其宿主CD276基因的表达降低,进一步提示宿主CD276可能为血吸虫mir-125b靶基因;荧光定量PCR分析表明感染日本血吸虫后的不同感染时期,小鼠淋巴细胞、肝脏及脾脏中的CD276的表达水平显著低于对照组(P0.01),进一步表明虫源性mir-125b在体内影响宿主CD276基因的表达。日本血吸虫mir-125b可能靶向调控宿主CD276基因的表达,在虫体与宿主互作中发挥重要调控功能。
[Abstract]:The host CD276 is the target gene of the mir-125b of Schistosoma japonicum by the luciferase experiment, and the expression of the host CD276 gene in different tissues after the infection of the schistosome is analyzed. The nucleic acid sequence interacting with the CD276 was amplified by PCR and the target fragment was cloned downstream of the expression luciferase reporter plasmid. The recombinant plasmid was transfected into the HEK293T cell to detect the luciferase reporter gene expression. The expression of the target gene CD276 was detected by the fluorescence quantitative PCR technique, and the expression of CD276 in different tissues of the mice was detected. Luciferase experiments show that the expression of the luciferase reporter gene of the transfected mir-125b mimomic and the recombinant plasmid is significantly reduced (P0.01), suggesting that the mir-125b can interact with the relevant region of the CD276 to inhibit the expression of the reporter gene; The expression of the host CD276 gene was reduced after the mouse macrophages were transfected with the schistosome mir-125b momic, and the host CD276 could be the target gene of the mir-125b of the SCHISTOSOMA. The fluorescence quantitative PCR analysis showed that the mouse lymphocytes, The expression of CD276 in the liver and spleen was significantly lower than that in the control group (P0.01), and the expression of the host CD276 gene in the host was further indicated. The Schistosoma japonicum mir-125b may target the expression of the host CD276 gene, and play an important role in the interaction of the worm and the host.
【作者单位】： 天津农学院;中国农业科学院上海兽医研究所农业部动物寄生虫学重点实验室;
【基金】：国家自然科学基金(31502056和31672550) 中国农业科学院科技创新工程
【分类号】：S852.7
[Abstract]:The host CD276 is the target gene of the mir-125b of Schistosoma japonicum by the luciferase experiment, and the expression of the host CD276 gene in different tissues after the infection of the schistosome is analyzed. The nucleic acid sequence interacting with the CD276 was amplified by PCR and the target fragment was cloned downstream of the expression luciferase reporter plasmid. The recombinant plasmid was transfected into the HEK293T cell to detect the luciferase reporter gene expression. The expression of the target gene CD276 was detected by the fluorescence quantitative PCR technique, and the expression of CD276 in different tissues of the mice was detected. Luciferase experiments show that the expression of the luciferase reporter gene of the transfected mir-125b mimomic and the recombinant plasmid is significantly reduced (P0.01), suggesting that the mir-125b can interact with the relevant region of the CD276 to inhibit the expression of the reporter gene; The expression of the host CD276 gene was reduced after the mouse macrophages were transfected with the schistosome mir-125b momic, and the host CD276 could be the target gene of the mir-125b of the SCHISTOSOMA. The fluorescence quantitative PCR analysis showed that the mouse lymphocytes, The expression of CD276 in the liver and spleen was significantly lower than that in the control group (P0.01), and the expression of the host CD276 gene in the host was further indicated. The Schistosoma japonicum mir-125b may target the expression of the host CD276 gene, and play an important role in the interaction of the worm and the host.
【作者单位】： 天津农学院;中国农业科学院上海兽医研究所农业部动物寄生虫学重点实验室;
【基金】：国家自然科学基金(31502056和31672550) 中国农业科学院科技创新工程
【分类号】：S852.7
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