Gene expression in suppressor mutants.

<p>(<b>A</b>) Western blot analysis of FPA protein levels. Three independent <i>clf-50 fpa-10</i> samples were processed. Note that no protein is detected in the null <i>fpa-7</i> control, whereas in extracts from a <i>35S::FPA-YPA</i> transgenic line a larger product corresponding to the <i>FPA-YFP</i> fusion protein is detected, confirming the specificity of the antibody for FPA. No FPA protein is detectable in <i>fpa-10</i> extracts, indicating that <i>fpa-10</i> is likely a null allele. (<b>B</b>) Real time PCR analysis of <i>FLC</i> expression. (<b>C</b>) Real time PCR analysis of <i>AG</i> expression, showing high <i>AG</i> expression in suppressor mutants. (<b>D</b>) Western blot analysis of AG protein expression. The AG antibody detects two proteins of about 29 kDa that are specific for AG, the smaller band possibly representing a truncated product or spurious translation initiation event (Riechmann et al., 1999). AG protein is strongly detected in wild type flowers but not in leaves. Weak expression is found in <i>clf-50</i> and <i>clf-50 fpa-10</i> leaves. (<b>E</b>) Real time PCR analysis of <i>SEP3</i> expression. (<b>F</b>) Histochemical staining of GUS reporter gene activity. <i>SEP3::GUS</i> is not expressed in wild type leaves but shows weak expression in vasculature of <i>clf-81</i> leaves (enlarged in inset). (<b>G</b>) Real time PCR analysis of <i>SEP2</i> expression. (<b>H</b>) Real time PCR analysis of <i>FT</i> expression (<b>I</b>) Real time PCR analysis of <i>SEP3</i> expression. Error bars in real time PCR experiments represent standard error of mean of three independent samples (biological replicates). Expression was normalised relative to the <i>EiF4A</i> gene, and is expressed relative to expression in wild type. In <b>B</b>, <b>C</b>, <b>E</b>, <b>G</b>, <b>H</b> whole seedlings less roots of 20 day old short day grown seedlings were used. In <b>I</b> rosette leaves of long day plants at 21 days were used.