Tissue Culture Responses of Elite Kenyan Maize inbred lines Via Somatic embryogenesis

Abstract

While conventional maize breeding has played a role in the development of improved crop varieties, it has the limitation of taking too long before achievement of an improved product; furthermore, it has failed to provide crop varieties with effective resistance to some diseases and insect pests such as larger grain borer. One of the possible promising options of developing maize varieties with resistance to larger grain borer is through genetic engineering (maize transformation). A key prerequisite to the transformation of maize is the establishment of sound regeneration system and identification of regenerable Kenyan maize genotypes. This investigation was undertaken in order to develop a plant regeneration protocol for some Kenyan maize inbred lines. Sixteen genotypes namely TL01, TL02, TL04, TL06, TL07, TH22, TL10, TL11, TL12, TL13, TL14, TL16, TL17, TL30, TL33 and TL34  of maize inbred lines were self-pollinated and immature Zygotic embryos measuring 1-1.5 mm were harvested at different days post pollination (DAP).Embryos harvested on days (10, 15, 18, 21 and 24),  were used to induce callus for the purpose of in vitro regeneration on N6 medium containing 2mg/l 2,4D (Optimal titrations), 2% sucrose and  supplemented with 2.88g L-proline, 0.2g casein hydrolysate devoid of silver nitrate, and   incubated in the dark at 25oC. All genotypes induced callus at an efficiency rate of 80%; six genotypes TL01, TL13, TL14, TL17, TH22 and TL33 produced somatic embryos in maturation medium containing 1mg/l NAA with 6% sucrose. The embryogenic calli produced shoots in MSO medium when incubated in full light at 28oC and a photoperiod of 16hours of light and 8 hours of darkness. This gave regeneration potential of inbred lines studied at 37.5%; of the six genotypes that regenerated, four genotypes produced seeds (TL01, TL13, TL14, and TL33). A comparison of the morphological characteristics between the original seeds R0 and the seeds from regenerated plants R1, showed no remarkable differences. TL01 produced one seed, while TL14 produced five seeds from the tassel, TL13 and TL33 produced seeds from the cobs. On planting R1 seeds alongside the Ro seeds the plants from the two seed types were normal in appearance indicating that, the regeneration system of Kenyan inbred lines was successfully completed. This is the first developed regeneration protocol for inbred lines in Sub- Saharan Africa which can be used for maize transformation