Cyclic Somatic Embryogenesis and Plant Regeneration in Cassava

With a four-step procedure, using the cassava clone M. Col 22, a cyclic system of somatic embryogenesis was developed. Primary embryos were induced in step 1 (2,4-D dependent) on young leaf lobe explants. Primary embryos were isolated in step 2 (BAP dependent) and were used as starting explant for secondary embryogenesis (steps 1 and 2 repeated). In this way, repetitive embryogenic cultures were established. It was shown that in all successive cycles embryo explants produced significantly more embryos than leaf lobe explants. According to their shape embryos were classified in two development stages; torpedo shaped embryos and germinated embryos. Germinated embryos produce more than three times as many embryos than torpedo shaped embryos. A third step in the embryogenic process was the development of shoots. The shoot conversion of embryos was increased if benzylamino purine (1 mg 1-1) was added. In that case more than half of the isolated embryos developed into normal shoots. The shoot conversion rate of germinated embryos was higher than that of torpedo shaped embryos. Embryos were kept in culture for 9 months to give six cycles. The production of embryos from the sixth cycle was as efficient as that from the second cycle. The shoot conversion rate of embryos from the fourth cycle was comparable with that from the second cycle. Rooting of the embryo-derived shoots (step 4 of the procedure) occurred on a medium without BAP. More than 100 regenerants obtained from fourth cycle embryos were multiplied in vitro. They showed no phenotypical differences as compared to in vitro multiplied parental material.