Figure 2.

shRNA mediated selective knockdown of Npn-2 or Plexin-A2 mRNAs in the chick ventral spinal cord induces ectopic positioning of motor neurons. (a-f) Confocal micrographs of transverse vibratome sections (75 μm) of HH stage 24 embryo spinal cord 2 days after electroporation in the ventral neural tube. MNR2/Islet2 positive ectopic motor neurons (red; white arrows) are found in embryos electroporated with shRNA-EGFP vectors specific for Npn-2 (a,b) and Plexin-A2 (e,f), but not for Plexin-A1 (c,d). The presence of the shRNA vector is indicated by EGFP expression (green) such that MNR2/Islet2 positive motor neuron somata in ectopic positions are yellow (arrows in (a,e)). Bar = 150 μm. (g) Histogram showing percentage of HH stage 24 embryo sections containing dual labelled EGFP and MNR2/Islet2 positive ectopic motor neurons after ventral electroporations at HH stage 12–15 with shRNA-EGFP vectors targeting Npn-1, Npn-2, Plexin-A1, Plexin-A2 and Plexin-A4, or EGFP control vector. Only those shRNA constructs targeting Npn-2 and Plexin-A2 induced ectopic positioning of motor neurons. ***P < 0.001; two-tailed t-test.