|Poly-A+ RNA Preparation Using Proteinase K and SDS |
NOTE: All solutions should be autoclaved or treated with 0.1% DEPC. Homogenizer probe should be treated with 0.5M NaOH for about 5 min, then rinsed with sterile H20 and STE. Econo column should be washed with 0.5M NaOH then several aliquots sterile H20.
This method can also be used for solid tissues. There is no hard rule for how much material to use, but since even a small piece of tissue is equivalent to a large number of cells, it is advisable to use a large volume of STE/proteinase K/SDS. The procedure works well for adult brain, mouse embryos and cell lines but not very well for other adult tissues. Acid phenol is the best technique for the latter.
|Disclaimer: The following set of protocols were contributed by various members of our lab (past and present): Christine Andrews, Fiona Christensen, Neil Della, Ross Dickins, Debbie Donald, Andrew Holloway, Gary Hime, Colin House, Yinling Hu, Rachael Parkinson, Nadia Traficante, Hannah Robertson, Ping Fu and Dennis Wang. Special thanks to Vicki Hammond, Frank Kontgen and Maria Murphy, who contributed many of the ES cell protocols. Sections dealing with Photomicroscopy, Polyclonal and Monoclonal Antibody Production were provided by members of Gerry Rubin's Laboratory (Berkeley). Any comments in the methods (technical errors etc.) E-mail: firstname.lastname@example.org |
David Bowtell PMCI October 1998