实时荧光定量PCR检测大鼠组织Angptl3表达的标准曲线构建

Construction of the standard curve for Real-time PCR detecting the expression of Angptl3 in rats

目的:采用实时荧光定量PCR(染料法)建立检测SD大鼠组织血管生成素样蛋白3(Angiopoietin-related protein3,Angptl3)表达量的标准曲线。方法:以SD大鼠肝组织总RNA为模板,通过RT-PCR获得目的DNA片段,用T-A克隆方法将其装载到质粒pUCm-T上,经过转化、质粒提取获得标准品,并以此为模板梯度稀释进行Real-time PCR反应,建立该方法的标准曲线。结果:Angptl3标准曲线相关系数为0.9976,相关性较好;斜率为0.8623,扩增效率适中。结论:成功构建了用Real-time PCR方法检测SD大鼠肝组织Angptl3表达量的标准曲线。

Objective： To construct the standard carve for real-time PCR detecting the tissue expression level of Angpt13 in SD rat. Methods：The total RNA of the SD rat＇s liver was the template.We got lots of the Angptl3 partial DNA sequence by RT-PCR.The products were cloned into the pUCm-T vector, Then we diluted the solution of plasmid and used real-time PCR （SYBR Green I ） to construct the stan-dard curve. Results：The correlation coefficient of the standard curve was 0.997 6. The reaction efficiency was 0.862 3. The correlation and the amplify efficiency were good. Conclusion：It is successful to construct the standard curve for real-time PCR detecting the tissue expression level of Angpt13 in SD rat.