Optimising Somatic Embryos Formation in Coffea arabica Cultivar Ruiru 11 Using Temporary Immersion Systems In Kenya

Abstract

Coffea arabica, F1 hybrid variety, Ruiru 11 is a highly sought-after crop in Kenya due to its alleviated resistance to Coffee Berry Disease and Coffee Leaf Rust coupled with high yield capacity and good cup quality. Access to the variety’s planting materials is limited due to challenges with difficulty in propagation using conventional methods of seed and vegetative propagation; and somatic embryogenesis is regarded as a suitable alternative propagation method. Temporary Immersion Systems are automated systems where cultures are submerged in liquid nutrient media using negative pressure for a specified immersion time then released. Temporary Immersion Systems are ideal for embryo induction and embryo maturation for C. arabica hybrids. Therefore, the current study aimed to establish an induction protocol in F1 composite hybrid Ruiru 11. The current study investigated the effects of genotype and plant growth regulators, auxins and cytokinins, on induction of embryogenic callus and embryo proliferation and maturation in two composite genotypes of C. arabica L. F1 hybrid variety Ruiru 11 -Code 71 and Code 93. Leaf explants from the F1 hybrid were cultured on half-strength Murashige and Skoog (MS) media supplemented with varied concentrations and combinations of plant growth regulators. Callus formation was evaluated weekly until the 60th day. Genotypic effects were assessed based on the difference on callus induction rates, callus proliferation and embryo induction and proliferation. The genotypes tested showed highest callus induction rates at 88% (Code 71) and 100% (Code 93) with respect to the formation of embryogenic calli. Highest fresh weight was obtained at 0.973 ± 0.011 g in Code 71 and 0.649 ± 0.03 g in Code 93 in MS media supplemented with 2,4-D + BAP (0.53 + 0.11 µM). Highest number of embryo induced was obtained from inoculum callus density 0.005 g/cm3 which generated 94.222 (Code 93) and 76.333 (Code 71) after two months in Temporary Immersion Systems (1-L RITA®) immersed for 3 minutes every 6 hours. As for callus proliferation in solid media (conventional method), Code 71 induced highest embryos at 15.11 in 84 days and in Code 93 at 56.333 after 91 days in 2,4-D at 0.33 µM and 0.44 µM respectively. Optimization achieved in Temporary Immersion Systems compared to conventional process based on time efficiency, embryo yield and resource use. TIS induced highest embryos at 94.222 after 170 days with elimination of gelrite which reduces cost of embryo production against conventional method induced highest embryos (56.333) after 162 days using solid media supplemented with gelrite. The study opens new prospects for Coffee Research Institute to adopt Temporary immersion systems for improved propagation of Ruiru 11 Coffee arabica hybrids for mass production to cater for increasing demand for coffee seedlings.