Total RNA isolation with RNA clean

Contributed by Dr. Alexei Gratchev

One of the best examples of phenol/guanidin-tiocyanate solutions for RNA isolation. The procedure is similar for all such reagents. The cells are lysed with the reagent on the plate or as a pellet, and then chlorophorm is added to extract the organic phase. Aqueous phase is collected and RNA is precipitated with ethanol. Pellet is washed and dissolved with water. With this method it is very easy to obtain highly concentrated RNA solution (up to 5ug/ul) However part of this concentration comes from tRNA which is efficiently precipitated, Therefore and impression that this method gives better RNA yeld than the previous one can be incorrect. Another problem I observed while using this kit is that usage of 1.5-2.0 ml tubes is the most efficient, but it results in dramtic increase of pipetting if big samples have to be processed. As soon as the volume of your lisate becomes more than 3 ml it's better to use another method. The time required for one preparation is similar to that of previous kit. You have to provide RNase free reagents. Therefore this method can be recommended for the labs where RNA work is good established and for small scale preparations when high concentration of RNA is required (for example for Northern blot). The protocol is provided with the reagent, but it seems to me a little to short and requires some experience.

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