Abstract: 【Objective】 The treatment of tillering node with colchicine in a pump solution has long been widely used for chromosome doubling of haploid wheat plants before transplanting. However, this method has some disadvantages, for example, complicated manipulation and environmental or container pollution. Moreover, colchicine is some toxic, and large amount of application is not cost effective. The purposes of this study were to develop a simple, safe and efficient method for chromosome doubling of wheat haploid plants, and to evaluate a chemical that is suitable for the replacement of colchicine for chromosome doubling in wheat. 【Method】 The haploid wheat plants were obtained by anther culture for wheat line Fielder, and by crossing with the maize inbred line Zheng58 for wheat cultivars/lines Kenong199, Xinchun9, Chinese Spring and CB037 via chromosome elimination. Wheat line Chinese Spring was also crossed with rye line Gansu Heimai to produce wheat-rye double haploid plants. Chromosome doubling of different wheat haploid plants was conducted using the colchicine solutions at the concentrations of 0, 5, 10, and 20 mmol·L^-1 through tiller node adding after transplanting, leaf painting after transplanting, and medium surface adding before transplanting. Additionally, the supplement of amiprophos-methyl, trifluralin, and propyzamide at the concentrations of 0, 30, 60 and 120 μmol·L^-1 on medium surface adding was used to double chromosomes of the wheat haploid plants and the wheat-rye double haploid plants. The efficiency of chromosome doubling by different treatment methods was compared and the optimal concentrations of different chemicals were determined. 【Result】 Results indicated that the application of different concentrations of colchicine (0, 5, 10, and 20 mmol·L^-1) by tiller node adding couldn’t be able to double the wheat haploid plants from the cross of wheat and maize, so it is not useful in chromosome doubling of wheat haploid plants. A doubling frequency of 7.7% was found when 10 mmol·L^-1 colchicine was applied by leaf painting for the wheat haploid plants from maize pollen induction at jointing stage, but no haploid plant was doubled with other three concentrations of colchicine solutions, suggesting that this treatment method was also not useful for the doubling of the wheat haploid plants. The frequencies of chromosome doubling for the wheat haploid plants generated from anther culture treated with the four colchicine concentrations by medium surface adding were 26.7%, 42.9%, 73.3%, and 85.7%, respectively. It is demonstrated that the supplement of colchicine solution on medium surface adding resulted in the highest efficiency of chromosome doubling for the wheat haploid plants and the optimal concentration is at least 20 mmol·L^-1. The frequency of chromosome doubling for the haploid plants generated from the cross of wheat and maize by medium surface adding of propyzamide at the concentrations of 0, 30, 60, and 20 μmol·L^-1 was 0, 0-57.1%, 28.6%-75.0%, and 0-100%. However, the other two herbicides used by this method were not effective in chromosome doubling. The frequency of chromosome doubling by medium surface adding of 120 μmol·L^-1 propyzamide for the wheat-rye double haploid plants was 9.0%. No seed was produced by the other three concentrations of propyzamide and the other three agents (amiprophos-methyl, trifluralin, and colchicine). 【Conclusion】 It is concluded that propyzamide at the concentration of 60-120 μmol·L^-1 produced promising results of chromosome doubling for the wheat haploid plants. The medium surface adding of colchicine and propyzamide at appropriate concentrations is effective and easily adopted to double chromosomes of wheat haploid plants.

Key words: wheat, haploid plants, distant hybridization, anther culture, doubling agents