三种不同方法固定的石蜡切片中RNA的分析

Analysis of RNA in Three Different Solvents Fixed Paraffin Embedded Slices

研究在 10 %中性福尔马林、丙酮、甲醇 氯仿 冰醋酸 3种方法固定的石蜡切片中提取RNA的质量和数量 .取 2 5 0g体重的Wistar大鼠的肾脏 ,分别采用 10 %中性福尔马林、丙酮、甲醇 氯仿 冰醋酸 3种方法固定 ,石蜡包埋 ,H E染色 ;采用RNA裂解液、TRIZOL试剂 2种方法提取切片RNA ,逆转录为cDNA ,采用普通PCR和SYBRGREEN 1定量PCR分析RNA质量和数量 .结果表明 ,3种固定方法都可保持组织良好的结构和形态 ;采用 2种提取方法 ,均可经RT PCR扩增出 180bp大鼠磷酸甘油醛脱氢酶 (G3PDH)、5 6 5bpβ肌动蛋白 (β actin)、10 0bp纤溶酶系活化剂抑制物 1(PAI 1) ;但采用RNA裂解液时 ,比TRIZOL试剂可提取更多的RNA .

For qualitative and quantitative analysis of RNA in 10% formalin, acetone or Methacarn (methanol chloroform acetic acid) fixed paraffin slices, the kidneys of 6 Wistar rats with 250 g body weight each were used. After fixed by formalin, acetone or Methacarn paraffin embedded and H E stained, the RNA was isolated by different methods such as RNA lysis buffer and TRIZOL reagent. The quality and quantity of RNA was analyzed by RT PCR and quantitative PCR. It was shown that the morphology of tissue was preserved well and 181 bp rat glyceraldehyde 3 phosphate (G3PDH) gene, 565 bp beta actin gene and 100 bp plasminogen activator inhibitor 1(PAI 1) gene could be detected by RT PCR in three different solvent fixed slices with two isolation methods. However, RNA lysis buffer could be considered to be better for isolating more RNA than TRIZOL in three different solvents fixed slices.