Somatic embryogenesis and genetic uniformity of regenerated cassava plants from low-temperature preserved secondary somatic cotyledons

Abstract

Low plant regeneration and transformation frequencies in cassava have been attributed to recycling of cassava secondary somatic embryos for long periods. This study examined the suitability of low-temperature storage of dehydrated cassava secondary somatic embryos as an alternative short-term conservation approach to recycling. The study included two experiments: in the first one, somatic embryogenesis from cotyledons of dehydrated secondary somatic embryos stored at six temperatures for 4, 8 and 12 months was established; and in the second, plant regeneration via shoot organogenesis using somatic cotyledon explants obtained in the first experiment was examined. The growth, photosynthetic pigment contents and genetic stability of the regenerated plants were assessed. At each storage temperature, the survival of explants, somatic embryogenesis, shoot induction and elongation decreased with increases in storage period. At 4 months after storage (MAS), explant survival, frequency of somatic embryogenesis, shoot induction, elongation and rooting had the best responses at 16EC and 20EC. Similarly, the growth and photosynthetic pigment contents of plants regenerated from somatic embryos preserved at 16EC and 20EC recorded the best results at 4 MAS. However, at 8 and 12 MAS, the best explant survival, somatic embryogenesis, shoot induction and elongation were recorded at 16EC. Eight RAPD primers produced a total of 56 distinct and scorable bands, ranging from 250 bp to 3000 bp in size. An random amplified DNA (RAPD) analysis showed a uniform banding profile among regenerants and to that of the mother plant, indicating no genetic variation among regenerants and between the regenerants and the mother plant. The data presented here suggest that cassava secondary somatic embryos dehydrated to 50% moisture content should be preserved at 16-20EC for 4 months; for 8 and 12 months storage, the embryos should be stored at 16EC. No genetic alteration was detected among the regenerants. Therefore, the method is a suitable means for short-term conservation of cassava secondary somatic embryos.

Somatic Embryogenesis and Genetic Uniformity of Cassava Plants Regenerated from Secondary Somatic Cotyledons Preserved in Osmotic Agents

“…Isolated secondary embryos were transferred to fresh BM supplemented with 2% mannitol, 2% sorbitol, 4% mannitol, 4% sorbitol, 2% mannitol + 2% sorbitol, 2% mannitol + 4% sorbitol, 4% mannitol and 2% sorbitol and a control (free of osmotic agents) for 8 and 16 months in the dark at 16 0 C. Preliminary studies have shown that storage at 16 0 C was the most suitable low temperature for a short-term preservation of cassava secondary somatic embryo (Opabode et al 2015).…”

“…The molecular technique of RAPD was used to determine the genetic stability of the regenerated plants as described by Opabode et al (2015). A total of eight random decamer primers (Operon Tech, Alameda, USA) from B, C, D, and E series (OPB06, OPB08, OPB12, OPC01, OPC02, OPC05, OPC06, OPE19) were used for RAPD analysis.…”