Doubled Haploid Laboratory Protocol for Wheat Using Wheat–Maize Wide Hybridization

Abstract

In traditional wheat breeding, the uniformity of lines derived from a breeding population is obtained by repeated selfing from the F1 which takes several generations to reach homozygosity in loci controlling traits of interest. Using doubled haploid technology, however, it is possible to attain 100% homozygosity at all loci in a single generation and completely homogeneous breeding lines can be obtained in 1–2 years. Thus, doubled haploid technology may significantly reduce cultivar development time. Two major methods for producing wheat doubled haploids are androgenesis (anther culture and microspore culture) and embryo culture using wheat–maize wide hybridization, the latter being the most effective and widely used method. The method of wide hybridization between wheat and maize is laborious but is widely successful for rapidly obtaining homozygous lines. This technique includes six major steps: emasculation of the wheat flower; pollination of the emasculated flower with maize pollen; hormone treatment; embryo rescue; haploid plant regeneration in tissue culture medium; and chromosome doubling. It has been observed that the efficiency of doubled haploid production depends on both maize and wheat genotypes, good plant health and proper greenhouse conditions (without disease, insects, or drought stress), and proper conduct of all procedures. Therefore, the procedures may need minor modification in order to produce higher numbers of embryos, haploid green plants, and doubled haploid plants.