GENELUTE DIRECT MRNA MINIPREP KIT

The Uses of GENELUTE DIRECT MRNA MINIPREP KIT

The purified bacterial genomic DNA is ready for downstream applications such as:

• restriction endonuclease digestions
• PCR
• Southern blots
• cloning

General Description

Sigma′s GenElute Mammalian Total RNA Miniprep Kit provides a simple and convenient way to isolate total RNA from mammalian cells and tissues. Protocols are provided for cells, tissues, and fibrous tissues. These protocols differ in their cell lysis and disruption conditions. Once the RNA is bound to the GenElute Binding Column, the purification procedure is the same for all starting materials. For fibrous tissues the kit must be used with proteinase K (Cat. No. P4850) to ensure effective cell disruption. The kit combines the advantages of a silica-based system with a microspin format and eliminates the need for cesium chloride gradients, alcohol precipitation, and hazardous organic compounds such as phenol and chloroform. Cells or tissues are lysed and homogenized in a buffer containing guanidine thiocyanate to ensure thorough
denaturation of macromolecules and inactivation of RNases. Addition of ethanol causes RNA to bind when the lysate is spun through a silica membrane in a microcentrifuge tube. After washing to remove contaminants, RNA is eluted in 50-100 μL of Elution Solution. Up to 150 μg of total RNA can be isolated in less than 30 minutes.

If all traces of DNA contamination must be eliminated, further treatment with DNase I is recommended. DNase I digestion can be performed while the RNA is bound to the GenElute Binding Column using the On-Column DNase I Digestion Set (DNASE10 and DNASE70). Alternatively, for more stringent removal of contaminating DNA, the final RNA preparation can be treated with Amplification Grade DNase I (AMPD1).