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Simple Isolation of RNA from Tissue and Cultured Cells


Easy RNA purification with integrated DNA-removal step

Karen Dolter Jeff Braman
Stratagene

The StrataPrep total RNA miniprep kit efficiently isolates high-quality total RNA from 10 5 to 10 7 cultured cells or 10 to 40 mg of tissue and includes an on-column DNA-removal step. Additionally, multiple samples can be processed simultaneously because of the easy-to-follow technique. The RNA isolation is performed within microspin cups, which eliminates the need for organic extraction or ethanol precipitation. The resulting total RNA is suitable for a wide variety of demanding molecular biology applications.

Fig.1

High-quality RNA is necessary for techniques such as cDNA synthesis, RT-PCR amplification, RNase protection assay, primer extension analysis, and Northern blotting. Stratagenes StrataPrep total RNA miniprep method takes advantage of the efficient denaturing properties of guanidine thiocyanate for cell lysis. The protein-denaturing ability of this chaotropic salt facilitates the isolation of intact RNA from tissue rich in ribonucleases. The method originally described1 has been simplified by using a silica-based fiber matrix in a microspin-cup format (Figure 1). The RNA is immobilized on the fiber matrix, allowing contaminant removal while avoiding organic extractions. In addition, no ethanol precipitation of the RNA is necessary. By treating the RNA sample directly on the fiber matrix with DNase, DNA contamination is reduced to undetectable levels. Once the purified RNA is eluted from the fiber matrix in a small volume of buffer or water, it is ready to use.

We showed that the kit is capable of high performance using
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