Protocol: high throughput silica-based purification of RNA from Arabidopsis seedlings in a 96-well format

Systems biology involves the study of molecules in context as part of a larger system or network rather than in isolation and the development of mathematical models of the particular system being studied. Both the wet (laboratory-based) and dry (computer-based) experiments are used to inform each other and together generate a greater understanding of the biological system [1,2]. Over the last few years there has been a movement towards a systems approach to studying biology with a concomitant year-on-year increase in publications to over 1,500 in 2009 [3] and this approach is being used in diverse fields such as stem cell differentiation (reviewed in [4]) and circadian rhythms in plants and animals (reviewed in [5]). As such systems-based approaches become increasingly more popular and incorporated in to a large variety of laboratories, there is a requirement for high throughput (HTP) experimental methods to be developed. As HTP procedures can be arduous for researchers to perform, automation is even more desirable especially when processing very large numbers of samples for an experiment.Historically, RNA extraction has been performed using organic solvents and phenol-chloroform [6]. Similar liquid-liquid extraction methods using commercially available reagents such as TRIZOL (Invitrogen/Life Technologies) or specifically targeted to more challenging high-polysaccharide-containing samples (such as plants and some Gram-negative bacteria) using CTAB (cetyltrimethylammonium bromide) have been developed [6]. These methods have been widely used, are relatively cheap and simple to perform and isolate large yields of high quality RNA. However, recently there have been concerns expressed over the use of TRIZOL with plant tissue [7,8]. An alternative is to use methods based on solid phase nucleic acid extraction (such as the selective binding of nucleic acid to silica matrices). These methods are commercially available as spin/vacuum-column kits and are convenient and effic