The E.Z.N.A.^® Plant RNA Midiprep Kit provides a convenient and rapid method for the isolation of total RNA from a variety of plant samples. Use of the HiBind^® system permits purification of high quality RNA from as much as 500 mg tissue. Plant RNA Kits are ideal for processing multiple plant samples in less than one hour. High quality plant total RNA can be reliably obtained from fresh or frozen (with liquid nitrogen) samples. Purified RNA from this kit has OD ratios that range from 1.8 to 2.0 and is suitable for a number of downstream applications.

Principle

The E.Z.N.A.^® Plant RNA Midiprep Kit combines the reversible nucleic acid-binding properties of HiBind^® matrix with the speed and versatility of spin column technology. HiBind™ matrix avidly, but reversibly, binds RNA under specific conditions allowing proteins and other contaminants to be removed. Nucleic acids are easily eluted with de-ionized water or low salt buffer. The kit also includes homogenizing columns to efficiently remove cell debris and simultaneously homogenize the sample. The method involves a simple and rapid precipitation step for the removal of polysaccharides, starches and phenolic compounds commonly found in plant tissues.

Procedure

Collect approximately 500 mg of finely ground plant material in a microfuge tube and immediately add 2.5 ml Buffer RB containing the appropriate amount of ß-mercaptoethanol and incubate. After incubation transfer lysate into a Homogenization Spin Column and centrifuge at >6,000 x g for 15 minutes at room temperature. After centrifugation, discard the homogenization column and add 0.5 volume of ethanol and mix by inverting tube. Apply the sample, including any precipitate that has formed, to a HiBind^® RNA spin column and centrifuge at 10,000 x g for 30 sec at room temperature. If highly pure RNA is required , an optional DNase I digestion reaction (Catalog Number E1091) can be conducted directly onto the surface of the HiBind^® RNA resin. RNA present on the column is subjected to several wash steps to remove proteins, polysaccharides and other contaminating compounds. Clean RNA is then eluted from the column with DEPC-treated water.

Applications

RNA purified using the E.Z.N.A.^® Plant RNA Midiprep Kit is suitable for the following applications :

• RT-PCR
• Northern Analysis
• Differential display
• Poly A⁺ RNA selection