Production of Doubled Haploids
Doubled haploids can be produced in vivo or in vitro. Haploid embryos are produced in vivo by parthenogenesis, pseudogamy, or chromosome elimination after wide crossing. The haploid embryo is rescued, cultured, and chromosome-doubling produces doubled haploids. The in vitro methods include gynogenesis (ovary and flower culture) and androgenesis (anther and microspore culture). Androgenesis is preferred method. Another method of producing the haploids is wide crossing. In barley, haploids can be produced by wide crossing with the related species Hordeum bulbosum, fertilization is effected, but during the early stages of seed development the H. bulbosum chromosomes are eliminated leaving a haploid embryo. In tobacco (Nicotiana tabacum), wide crossing with Nicotiana africana is widely used. When N. africana is used to pollinate N. tabacum, 0.25 to 1.42 percent of the progeny survive and can readily be identified as either F1 hybrids or maternal haploids. Although, these percentages appear small, the vast yield of tiny seeds and the early death of most seedlings provide significant numbers of viable hybrids and haploids in relatively small soil containers. This method of interspecific pollination serves as a practical way of producing seed-derived haploids of N. tabacum, either as an alternative method or complementary method to anther culture.
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