Figure 2.

Microarray detection of cellular RNAs, including ncRNAs, that associate with the bacterial Sm-like protein, Hfq, and the La homologous protein (Lhp1p) of yeast (S.cerevisiae), respectively (77,87). Left panel: RNAs are co-immunoprecipitated from E.coli cell extracts with anti-Hfq antibodies, purified and hybridized to high-density microarrays that carry DNA oligonucleotides covering the entire E.coli genome. RNAs that hybridized to probes on the arrays are detected with an antibody that specifically sees DNA:RNA hybrids. Subsequently, such signals are detected as indicated and subtracted from those obtained in a control experiment in which cell extracts were incubated with pre-immune sera, that is no immunoprecipitation of Hfq. Right panel: cell extracts from either wild-type yeast or yeast cells that express epitope (myc)-tagged Lhp1p are incubated with an anti-myc antibody, RNA is extracted from immunoprecipitates and reverse transcribed. The two obtained cDNAs are labelled with different fluorescent dyes (Cy3, red; Cy5, green), mixed and hybridized to yeast whole-genome microarrays. Spots that yield red signals indicate that the corresponding RNA was enriched in Lhp1p-myc immunoprecipitates.