Culture of freshly isolated wheat (Triticum aestivum L.) microspores treated with inducer chemicals
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Microspores were isolated from wheat (Triticum aestivum L.) spikes by means of a micro-blender immediately following their removal from the donor plants. Isolated microspores were then subjected to a chemical treatment consisting of 0.18 mM 2-hydroxynicotinic acid, minerals and 9% maltose in the dark at 25°C for 38–52 h. Following purification via filtration and gradient centrifugation on 21% maltose, the microspores were cultured in the presence of excised ovaries in liquid medium at 27°C. Embryoid yield, percentage of green plants, and the frequency of spontaneously doubled haploids ranged from 360 to 4,914 embryoids per spike, 15% to 95%, and 39% to 78%, respectively. Other compounds that were effective in maintaining viability and triggering microspore embryogenesis were benzotriazole-5-carboxylic acid and violuric acid monohydrate. This system has proven to be highly efficient for producing doubled haploids over a range of genotypes.
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