Analysis of RNA by Northern and Slot Blot Hybridization
Analysis of RNA by Northern and Slot Blot Hybridization
复制标题
通过 Northern 杂交和狭缝印迹杂交分析 RNA
DOI：
10.1002/0471142727.mb0409s37
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发表时间：
1997
影响因子：
--
通讯作者：
Karol Mackey
中科院分区：
文献类型：
--
作者：
T. Brown;Karol Mackey
RNA制剂中的特定序列可以通过印迹和杂交分析来检测，使用的技术与最初用于DNA的技术非常相似。分离的RNA从琼脂糖凝胶转移到膜载体（northern blotting）；未分离的RNA通过槽或点印迹固定。用标记的DNA或RNA探针进行杂交分析。Northern blotting与Southern blotting在最初的凝胶分离步骤中有很大的不同。由于它们是单链的，大多数rna能够通过分子内碱基配对形成二级结构，因此如果要获得良好的分离，就必须在变性条件下进行电泳。通过在凝胶和装载缓冲液中添加甲醛或在装载前用乙二醛和二甲基亚砜（DMSO）处理RNA来实现变性。描述了琼脂糖甲醛凝胶中分离的RNA的印迹和杂交。备选方案描述了乙二醛/DMSO法变性凝胶电泳和RNA样品的槽印迹杂交。从印迹中剥离杂交探针可以在三种不同的条件下完成；这些方法概述在a。
Specific sequences in RNA preparations can be detected by blotting and hybridization analysis using techniques very similar to those originally developed for DNA. Fractionated RNA is transferred from an agarose gel to a membrane support (northern blotting); unfractionated RNA is immobilized by slot or dot blotting. The resulting blots are studied by hybridization analysis with labeled DNA or RNA probes. Northern blotting differs from Southern blotting largely in the initial gel fractionation step. Because they are single‐stranded, most RNAs are able to form secondary structures by intramolecular base pairing and must therefore be electrophoresed under denaturing conditions if good separations are to be obtained. Denaturation is achieved either by adding formaldehyde to the gel and loading buffers or by treating the RNA with glyoxal and dimethyl sulfoxide (DMSO) prior to loading. The describes blotting and hybridization of RNA fractionated in an agarose‐formaldehyde gel. Alternate protocols describe the glyoxal/DMSO method for denaturing gel electrophoresis and slot‐blot hybridization of RNA samples. Stripping hybridization probes from blots can be done under three different sets of conditions; these methods are outlined in a .
DNA 从聚丙烯酰胺或琼脂糖凝胶到硝化纤维素的定量电泳转移。
影响因子：
2.9
作者：
Smith,MR;Devine,CS;Cohn,SM;Lieberman,MW
通讯作者：
Lieberman,MW
影响因子：
2.9
作者：
CHOMCZYNSKI, P
通讯作者：
CHOMCZYNSKI, P