通过S1核酸酶消化及吸附到DEAE-纤维素滤膜上对DNA-RNA杂交体进行分析。
Assay of DNA-RNA hybrids by S1 nuclease digestion and adsorption to DEAE-cellulose filters.
作者信息
Maxwell I H, Van Ness J, Hahn W E
出版信息
Nucleic Acids Res. 1978 Jun;5(6):2033-8. doi: 10.1093/nar/5.6.2033.
Abstract
A fast and accurate assay procedure for DNA-RNA hybrids is described in which exhaustive digestion of unhybridized DNA with S1 nuclease is followed by binding of hybrids to filter discs of DEAE-cellulose. The digested DNA can be efficiently washed from the filters so that background levels of 0.1-0.2% of input tracer DNA can be achieved, in contrast to the much higher (approximately 1-5%) backgrounds obtained using TCA precipitation procedures. Short duplexes, as small as 36 nucleotides in length, which are inefficiently bound to hydroxyapatite, are quantitatively bound to the DEAE-cellulose filters.
摘要
本文描述了一种用于DNA-RNA杂交体的快速且准确的检测方法。该方法先用S1核酸酶彻底消化未杂交的DNA，然后将杂交体与DEAE-纤维素滤膜结合。消化后的DNA能有效地从滤膜上洗去，从而可使背景水平达到输入示踪DNA的0.1-0.2%，相比之下，使用三氯乙酸沉淀法获得的背景水平要高得多（约1-5%）。短至36个核苷酸长度的短双链体，其与羟基磷灰石的结合效率较低，但能定量结合到DEAE-纤维素滤膜上。
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