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In vitro technology has been advancing as a highly valuable pre-breeding tool for achieving better results in germplasm preservation, distant hybridization and genetic transformation approaches. Furtheremore, it has been gaining a growing importance in direct applications in conventional line selection by facilitating asexual multiplication, inducing variations, undertaking site-directed mutagenesis and production of doubled haploids. Because of genetic difference, tissue specificity and culture environment (media) interactions, there are no universally applicable protocols of culture and regeneration pathways. It varies as tissues from different sources and species as well as varieties differ in their response and culture requirements. The challenges are more conspicuous with recalcitrant monocot species such as tef.

An in vitro culture protocol was established for direct regeneration of plantlets from unpollinated ovary cultures of four Ethiopian wheat varieties. Unpollinated ovaries were excised from durum wheat (Yerer and Ude varieties) and bread wheat (Simba and Galama varieties). Analysis of variance (ANOVA) has shown that genotypes, types of media, concentrations of 2,4-dichlorophenoxy acetic acid (2,4-D) and kinetin (KIN) and durations of cold pretreatment at 4°C significantly (P≤0.05) affected direct formation of embryonic tissues independently. Stage II of wheat spikes, MS medium containing 1 mg/l of each of 2,4-D and KIN and 15 days of cold pretreatment were found to be the best conditions for direct formation of embryonic tissues. The highest frequency of shoots were regenerated from the cultured embryonic tissues of Yerer (41.6% ) and Simba (41.3%) on medium containing 0.1 mg/l 2,4-D. From a total of 14,524 cultured unpollinated ovaries, 1,100 embryonic tissues (7.6 %) and 75 regenerants were obtained. The average percentage of embryonic tissues and regenerants were 9.0 and 1.1% from 3,444; 9.8 and 0.55% from 4,732; 5.6 and 0.17% from 2,988; 4.7 and 0.12% from 3,360 cultured unpollinated ovaries for varieties Yerer, Simba, Ude and Galama, respectively.Keywords: Embryonic tissues, Unpollinated ovaries, regenerants, wheat varietiesAfrican Journal of Biotechnology Vol. 12(39), pp. 5754-5760