Efficient isolation of microspores and the production of microspore-derived embryos from Brassica napus

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Abstract
Mechanical isolation of Brassica napus microspores from whole buds with a micro-blender enabled the rapid isolation of large numbers of microspores free from tissue and cellular debris. The procedure resulted in savings of time and labor and could be used independent of the size and number of buds to be examined. Between 700 and 1000 embryos per bud were obtained. Compared to anther removal there was no difference in embryo yields or quality. Microspore isolation under cool conditions and overnight incubation prior to plating improved the frequency of embryogenesis. Over 75% of the embryos developed into normal torpedo-stage structures if the medium was replenished during culture and the embryos placed on a gyratory shaker. Over 80% of the torpedo-shaped embryos would ultimately develop into plants. The implications of these techniques to genetic and physiological studies are discussed.