Isolation of BnCLE19 from microspore-derived embryos of B. napus by DD-RT-PCR. (A) Schematic presentation of the B. napus microspore culture system used to obtain a large amount of relatively synchronised embryos. Immature microspores at the late uni-cellular and early bi-cellular stages were isolated and cultured at different temperatures, leading to either embryogenesis (32 °C) or pollen maturation (18 °C). (B) A portion of the DD-RT-PCR auto-radiograph showing the expression of BnCLE19 (arrow) in B. napus embryos after 10 days (globular to heart-shaped stage) and 16 days (torpedo stage) of culture at 32 °C. BnCLE19 mRNA was not detectable in freshly isolated microspores (T=0), in microspores cultured for 8 h at 18 or 32 °C, or at 41 °C for 45 min, or in leaves.