【摘要】 目的对3种血浆微小RNA(miRNA)提取方法进行评价。方法运用国产、进口硅胶吸附柱法和TRIzol法分别提取20名志愿者血浆中的miRNA;利用TaqMan聚合酶链反应(PCR)检测提取物中的β-globin基因含量,观察提取物中RNA的纯度;利用反转录茎环引物及TaqMan-MGB探针q PCR检测提取物中miRNA-21含量,观察miRNA提取的效率。结果国产、进口硅胶吸附柱法及TRIzol法提取液中β-globin基因的Ct值分别为35.487±0.356、35.591±0.332和33.529±0.499,3种方法的Ct值差异有统计学意义(P<0.001);国产和进口硅胶吸附柱方法的β-globin基因Ct值均明显高于TRIzol法(P<0.000 1);而国产与进口硅胶吸附柱法β-globin基因Ct值差异无统计学意义(P=0.345 4)。国产、进口硅胶吸附柱法和TRIzol法miRNA-21的Ct值分别为26.527±0.158、26.653±0.201和28.169±0.385,3种方法的Ct值差异有统计学意义(P=0.000 3);国产和进口硅胶吸附柱法miRNA-21的Ct值均明显低于TRIzol法(P<0.000 1);国产硅胶吸附柱法miRNA-21的Ct值低于进口硅胶吸附柱法(P=0.033 7)。结论硅胶吸附柱法可有效提高miRNA的提取效率和纯度。国产硅胶吸附柱法完全可以取代进口硅胶吸附柱法。更多还原

【Abstract】 Objective To evaluate 3 methods of plasma micro RNA(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 TaqManMGB 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-globingene 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.更多还原