快速一步法抽提心肌组织细胞总RNA

RAPID ONE -STEP METHOD FOR ISOLATION OF TOTAL RNA FROM RAT MYOCARDIAL TISSIUS

用快速一步法（简称快速法）抽提心肌组织细胞总ＲＮＡ，产量达２．１９±０．２９μｇ·ｍｇ－１心肌组织，纯度指标（Ａ２６０／２８０）为１．９３±０．０３；各指标均高于ＡＧＰＣ法（Ｐ＜０．０１）。操作时间由４ｈ缩短至２．５ｈ。心肌组织细胞，总ＲＮＡ置普通琼脂糖凝胶上电泳，可见２８Ｓ，１８Ｓ和５Ｓ（５．８Ｓ）三条清晰的区带。Ｎｏｒｔｈｅｒｎ杂交证明该法抽提的总ＲＮＡ无ＤＮＡ污染，不降解。快速法可取代ＡＧＰＣ法。结果还提示，２８Ｓ与１８Ｓ比值不能用来判断纯化后ＲＮＡ的完整性。

NA isolation procedure by AGPC(Acid guanidium thiocyanate-phenol-chloroform)extrac-tion has not only consumed time and reagents,but also resulted in a lower yield of total RNA be-cause of loss of low molecular-weight RNA(such as 5S,5.8S RNA).A rapid one-step methodfor isolation of total RNA from rat mvocardial tissue is described in this report.The yield of totalRNA isolated by this method was 2.19 ± 0.29μg·mg-1 tissue;the ratio of A260/280(purityindex)was l.93±0.03.All of them were higher than those by AGPC method(P <0.0l)Gen-eral agarose gel resolution patterns of total RNA revealed three bands of 28S,18S,5.8S and 5SRNA. Northern blot analysis showed that total RNA were free of DNA and in an undegradedform. The procedure required little time and less reagents. It is considered that the rapid one-stepmethod is a useful alternative to the AGPC method for RNA isolation,The results also suggestthat the ratio of 28S/18S can not be used to judge the integrity of the purified RNA.