At the end of culture, viability was confirmed by observing the blood circulation of the yolk sac and the beating heart. Cultured embryos were quickly immersed in 0.7 ml TRIzol (Invitrogen, Carlsbad, CA) and homogenized for extracting total RNA for the RT-PCR and microarray processes (see microarray section, below), or fixed in 4% paraformaldehyde in PBS for the evaluation of the developmental status.
