关键词: 微小RNA, 核酸提取, 硅胶吸附柱, TRIzol法, TaqMan聚合酶链反应

Abstract:

Objective To evaluate 3 methods of plasma microRNA（miRNA） extraction. Methods Plasma miRNA of 20 volunteers was extracted by domestic and imported silica gel adsorption column methods and TRIzol method. The content of β-globin gene was determined by TaqMan polymerase chain reaction（PCR） to observe the purity of RNA. The miRNA-21 content was determined by reverse transcription stem loop primers and TaqMan-MGB probe PCR to observe the efficiency of miRNA extraction. Results The Ct values of β-globin gene extracted by domestic and imported silica gel adsorption column methods and TRIzol method were 35.487±0.356,35.591±0.332 and 33.529±0.499,respectively. The Ct values of β-globin gene by the 3 methods had statistical significance（P<0.001）. The Ct values of β-globin gene by domestic and imported silica gel adsorption column methods were higher than that by TRIzol method（P<0.000 1）. There was no statistical significance for the Ct values of β-globin gene between the 2 silica gel adsorption column methods（P=0.345 4）. The miRNA-21 Ct values of domestic and imported silica gel adsorption column methods and TRIzol method were 26.527±0.158,26.653±0.201 and 28.169±0.385,respectively. There was statistical significance among the 3 methods（P=0.000 3）. The miRNA-21 Ct values of domestic and imported silica gel adsorption column methods were lower than that of TRIzol method（P<0.000 1）. The miRNA-21 Ct value of domestic silica gel adsorption column method was lower than that of imported one（P=0.033 7）. Conclusions The silica gel adsorption column method for extracting miRNA can effectively improve the extraction efficiency and purity of miRNA,and the domestic silica gel adsorption column method can replace the imported one.

Key words: MicroRNA, Nucleic acid extraction, Silica gel adsorption column, TRIzol, TaqMan polymerase chain reaction