High Pure RNA Tissue Kit

The High Pure RNA Tissue Kit is designed for the purification of total, intact RNA from tissue samples, free of any contaminating DNA. Low to medium throughput RNA isolation. Nucleic acids bind to the surface of the glass fleece in the presence of a chaotropic salt (guanidine HCl). This allows the High Pure Filter Tube to specifically immobilize nucleic acids (both DNA and RNA) while they are freed of contaminants. For RNA isolation, the binding conditions can be optimized to ensure immobilization of all the RNA. High pure RNA isolation kit can rapidly isolate intact total RNA from a broad range of research sample materials, including cultured cells, mammalian blood, white blood cells (WBCs), yeast, and bacteria. It rapidly isolates total RNA from mammalian tissues such as mouse liver, spleen, lung, and heart.

Capacity: The High Pure Spin Filter Tubes hold up to 700 μL sample volume.
Sample Material: Solid tissue (e.g., mouse liver, spleen, lung, heart): 1 - 25 mg.

Note: Sample size depends on the method used to prepare the tissue; larger samples (>10 mg) should be processed via rotor-stator homogenization.

High Pure RNA Tissue Kit has been used:

• to extract total RNA from human cells for cDNA library generation
• to extract total RNA from cultured cells for real time PCR analysis
• to extract total RNAs from rat hippocampi
• to purify total RNA from cultured cells and human spinal cord
• to isolate total RNA of tissues collected from ventricle walls

The isolated RNA can be used in many downstream applications:,

• Northern blotting
• RACE
• primer extension
• differential display
• RNase protection assay
• in vitro translation

Tissue samples are disrupted and homogenized in the presence of a chaotropic salt (guanidine HCL) that inactivates RNases. The homogenate is then applied to the glass fiber fleece in a High Pure Spin Filter Tube. Under the buffer conditions used in the procedure, all nucleic acids bind to the glass fiber fleece, while contaminating substances (salts, proteins, and other cellular contaminants) do not. DNA in the preparation is digested with DNase I directly on the filter. Brief wash-and-spin steps readily remove the digested DNA fragments and other cellular contaminants. The remaining purified RNA is then eluted in a small volume of low-salt buffer.

High Pure Technology and Silica Adsorption Kits
For life science research only. Not for use in diagnostic procedures.

Figure 1: Comparison of different homogenization procedures. Mouse liver was homogenized using various procedures. Total RNA was purified from each of these homogenates with the High Pure RNA Tissue Kit. Aliquots of the isolated RNA were reverse transcribed with primers specific for a region of the GADH mRNA. cDNA products were analyzed via gel electrophoresis.

Lane 1: Homogenization: Ultra Turrax; yield: 1.9 μg/mg tissue; A 260/ A 280 : 2.0
Lane 2: Homogenization: motor-driven, disposable plastic pestle; yield: 3 μg/mg tissue; A 260 /A 280 : 2.0
Lane 3: Homogenization: mortar and pestle, 20 G needle; yield: 1.5 μg/mg tissue; A 260 /A 280 : 2.0
Lane 4: Homogenization: manual disposable plastic pestle, 20 G needle; yield: 1.8 μg/mg tissue; A 260 /A 280 : 2.0
Lane 5: Homogenization: manual disposable plastic pestle; yield: 3.4 μg/mg tissue; A 260 /A 280 : 2.0
Lane 6: Homogenization: bead-vortex; yield: 3 μg/mg tissue; A 260 /A 280 : 2.0
Lane 7: Molecular weight marker