To determine the expected number of differentially expressed “neighbouring” genes occurring by chance assuming that the knockdown has no effect on gene expression, we calculated the average number of genes in a 300Kb window around a randomly selected gene in the human and mouse genome. We calculated this to be 11.2 (human) and 11.8 (mouse). For simplicity, we will conservatively round this down to 11. Assuming that no genes are changing between the knockdown and control, using a nominal p-value, which has a uniform distribution under the null hypothesis (nothing effected), we would expect to see a difference called in 5% of cases at a p-value of 0.05. If we test one locus, which has on average 11 neighbours we would expect to identify 0.55 hits by chance (11 × 0.05=0.55). However, if we now test 12 loci we would expect to see 6.6 (12 × 0.55) knockdowns which appear to have an effect under the null hypothesis.
