Total RNA SAFEKIT
Isolation of Total RNA from Tissues and cells without the use of hazardous chemicals

BIO 101 introduces a kit for the isolation of RNA that requires no organic extraction solvents or guanidine salts. The procedure is characterized by its simplicity and the resulting RNA is undegraded, free of DNA and protein, and is unusually stable. As opposed to other methods involving chaotropic salts and or lithium salt precipitation, that may or may not eliminate DNA, the RNA SAFEKIT completely eliminates DNA by use of a catalytic method that is stable at room temperature. The method includes cell lysis in the presence of Solutions 1 and 2 containing RNA stabilizing chaotropes to solubilize proteins and RNA and enable most DNA and cellular debris to be precipitated. RNA and traces of DNA are precipitated from the clarified supernatant. The RNA is finally purified by newly developed proprietary procedures involving catalytic removal of DNA and RNA stabilization. The resulting RNA is application ready and because it contains no DNA it is especially suitable for RT-PCR and differential display, directly, without the need to DNase-treat.

The protocol, requiring 1/2 to 1 hour, in brief:

1. Homogenize 10 to 100 mg of a tissue or cell sample (by manual or automated mechanical means) with 800 µl of Solution 1 and 200 µl of Solution 2. Spin lysate in a microcentrifuge for 5 minutes to pellet cellular debris.
2. Transfer 800 µl to another tube, add an equal volume of isopropanol to precipitate RNA and spin to pellet RNA.
3. Dissolve RNA in 200 µl of Solution 3 and add 40 µl of Solution 4 matrix suspension, mix, spin, and transfer supernatant containing purified RNA to another tube.

RNA is ready to use without further manipulation.

Total RNA SAFEKIT
Cat. No.	Description	Size/Vol.
1008-200	Total RNA SAFEKIT	50 Preps (yield 100µg/prep)
Kit Contents
1008-201	Solution 1	45 ml
1008-202	Solution 2	12 ml
1008-203	Solution 3	12 ml
1008-204	Solution 4	2.5 ml
1007-205	10% Acetic Acid	0.5 ml
1009-206	Cell Lysis Solution	75 ml
1009-207	*Phenol Buffer	30 ml
1009-208	**Isoamyl Alcohol	2.2 ml
User supplies: *Phenol, **Chloroform

Total RNA prep from mouse liver (lane a) and brain tissue (lane b). using the Total RNA SAFEKIT.

RT-PCR with ß globin primers of total RNA isolated from mouse liver shown in lane a: Lane 1 was with RT and lane 2 was without RT, indicating that no DNA is present in the RNA prep using the RNA SAFEKIT.