<i>ppk23</i> expression is reduced in appendages of the <i>Poxn</i> mutant.

<p>(A) Real-time quantitative RT-PCR analysis of total RNA extracted from adult appendages (legs and wings) from a mixed sex population. Analysis compared <i>Poxn</i>^M22-B5, which do not have external chemosensory bristles, and <i>CyO</i> balanced siblings, which develop normal sensory bristles <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002587#pgen.1002587-Boll1" target="_blank">[54]</a>. For illustrative purposes, data are represented as relative expression fold differences in <i>Poxn</i> flies relative to controls <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002587#pgen.1002587-BenShahar3" target="_blank">[51]</a>. Each data point includes the relative expression of a gene in <i>Poxn</i> homozygous flies relative to balanced <i>Poxn/CyO</i> flies, which develop normal sensory system. <i>CheB42a</i> gene was used as positive controls for chemosensory specific genes expressed in appendages <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002587#pgen.1002587-BenShahar1" target="_blank">[16]</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002587#pgen.1002587-BenShahar2" target="_blank">[17]</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002587#pgen.1002587-Xu1" target="_blank">[55]</a>. Statistical analyses were performed on the ΔCt data as previously described <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002587#pgen.1002587-BenShahar1" target="_blank">[16]</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002587#pgen.1002587-BenShahar2" target="_blank">[17]</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002587#pgen.1002587-BenShahar3" target="_blank">[51]</a>. *, <i>p</i><0.05; **, <i>p</i><0.01 (n = 4 per genotype, one-tail paired <i>t</i>-test) (B) Northern blot analysis of <i>ppk23</i> spatial expression patterns. Lower panel shows ribosomal bands on the RNA gel, indicating equal sample loading. Using RT-PCR coupled with 5′ and 3′ RACE analyses on RNA extracted from male appendages, we were able to identify only a single <i>ppk23</i> transcript (black arrow; <i>ppk23</i>-RX; see also <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002587#pgen.1002587.s001" target="_blank">Figure S1A</a>). Numbers on right represent RNA size markers (bp). (C) <i>ppk23</i> promoter activity in forelegs of male and female flies (reporter was nuclear GFP). Scale bar represents 50 µm. (D) Quantitative data from (C) showing significantly fewer <i>ppk23</i>-positive cells in the female foreleg (Independent sample <i>t</i>-test; *, p<0.05; n = 10 for each bar).