Protocol of triticale (x Triticosecale Wittmack) microspore culture

Abstract

Triticale (x Triticosecale Wittmack) is a spontaneous synthetic amphiploid cereal that has been considerably improved through breeding. Triticale is currently grown on about 2 million hectares worldwide. After the initial period, intensive breeding research on triticale was started in the early 1950s by Arpad Kiss in Hungary. Nowadays, the centre of European triticale breeding is in Poland. Polish breeders released excellent varieties that have had a good performance, especially in Europe. Since the induction of the first triticale anther culture derived haploid plantlets by Wang et al. (1973), the method of anther culture has been essentially modified and improved. Theoretically, two in vitro cell and tissue culture methods are used to induce androgenesis: anther and microspore culture. The anther culture method seems to be more laborious. The blender isolation protocol provides a sufficient number of microspores to avoid the necessity for anther isolation. In cereals, microblending isolation was earlier established for barley (Olsen, 1991) and wheat (Ziauddin et al.,1992), confirming its potential. The present protocol describes an in vitro process for blender-isolated triticale microspores leading to doubled haploid lines from microspore derived embryo-like structures (ELS). Embryogenesis was observed in isolated triticale microspores. This phenomenon also occurs in microspore culture of other cereals. From the ELS green plants can be regenerated, but albinism is still a typical problem of triticale microspore culture. Depending on the genotype, more than half of the regenerants may be albino.