Puf proteins, RNA co-immunopurification. Saccharomyces cerevisiae

RNA-coimmunopurifications with TAP-tagged Puf proteins from Saccharomyces cereviseae. Untagged strain (BY4741) served as a control. Cells were grown to midlog phase and harvested by centrifugation. TAP-tagged Puf proteins were affinity purified from cell-free extracts with IgG sepharose and eluted with TEV protease. RNA was isolated from extract (=input)and from purified protein samples by phenol-chloroform extraction. RNA samples were reverse transcribed using a mixture of oligo-dT and random nonamer oligos in the presence of amino-allyl dUTP/ dNTP mixture. cDNAs were fluorescently labeled and hybridized on yeast DNA microarrays over night at 65 degrees. For a detailed procedure see http://microarray-pubs.stanford.edu/yeast_puf and also Gerber AP et al. PLoS Biology, 2004. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Keywords: Logical Set Overall design: Computed