Faithful transmission and expression of transgenes via somatic embryos of transgenic cassava plants at the sixth cycle of vegetative propagation

Abstract

The transmission of transgenes via somatic embryos from one vegetative cycle to another in cassava (Manihot esculenta Crantz) has not been well studied to date. This study examined somatic embryogenesis and regeneration from transgenic cassava plants expressing the β -glucuronidase gene (GUS) under the control of a cassava vein mosaic promoter (CsVMV) at the sixth cycle of vegetative propagation. Primary, secondary and cyclic somatic embryos were induced from an axillary bud and immature leaf lobe explants. Plantlet regeneration via shoot organogenesis was examined in the cotyledons of somatic embryos. Histochemical, polymerase chain reaction (PCR) and reverse transcriptase PCR analyzes were used to confirm the presence of the GUS gene in regenerated plants. The frequencies of primary, secondary and cyclic somatic embryos from immature leaf lobes were significantly greater (P < 0.05) than those of axillary buds. The transient expression of the GUS gene was detected in all types of somatic embryos from both axillary bud and immature leaf lobe explants. Shoot induction from cotyledons of somatic embryos produced from axillary buds was 27.0% greater than that of leaf lobes, with selection performed with the use of kanamycin. In regenerated transgenic plants (grown from somatic embryos obtained from immature leaf lobes), a non-uniform expression of GUS was observed in three chimeric lines assessed by histochemical and molecular analyzes. In regenerated transgenic plants obtained from somatic embryos from axillary buds, uniform and high expression of GUS was observed in one line in all tested tissues and in most cell types of leaves, stems, petioles, roots and tubers, thus showing the faithful transmission of the GUS transgene under the control of CsVMV promoter via a somatic embryo.