Title : Efficient production of haploid durum wheat plant via maize pollination
Abstract:
Even though bread wheat haploids are being routinely produced by wheat × maize crosses, an efficient protocol for haploid production has been difficult to achieve in durum wheat. The objective of this study was to analyse the influence of various experimental factors on the production of embryos and haploid plants in durum wheat crossed with maize. Four Tunisian durum wheat genotypes (female parent), consisting of two local cultivars (‘Jenah khotifa’ and ‘Biskri’) and two improved varieties (‘Karim’ and ‘Razzek’), were crossed with a maize genotype (Pioneer 37Y15) (male parent). After pollination, plant stems were either maintained in situ or cut 2-3 days after pollination near the base and kept in a solution of 2,4- dichlorophenoxyacetic acid (100 mg/l), sucrose (40 g/l) and sulphurous acid (8 ml/l). Twelve, 14, 16, 18, 20 and 22 days after pollination, embryos were excised from developed ovaries and cultured on either Murashige and Skoog (MS), MS/2 or Gamborg’s (B5) media. The effect of different concentrations (0, 25, 50, 75, 100, 125, 150 and 175 mg/l) of 2, 4-dichlorophenoxyacetic acid (2,4-D) and nitrate silver (AgNO3) in sucrose solution and the combination of these two treatments (100 mg/l of 2,4-D and 75 mg/l of AgNO3) were tested. Performance of durum wheat doubled haploids derived from durum wheat x maize crosses and their parents was evaluated on the basis of germination percentage and six morphological traits such as plant height, spikeless tillers/plant, spike tillers/plant, days from sowing to 50% ear emergence, kernels/spike and 1000-kernel weight.
The results showed that by cutting the plant stems after pollination, better results in terms of developed ovaries (65.55%), embryos (13.62%) and regenerated haploid plants (10.50%) were obtained than if in situ plants (46.65, 10.85 and 7.71%) were used. The optimal stage for embryo rescue was 18 days for haploid plant formation (14.4%) while B5 medium resulted in significantly better haploid plant (13.17%) production than MS and MS/2 (6.92, 10.87%) following embryo culture. The best results of embryo formation and haploid plants were obtained when the concentration of 2,4-D was 100 mg/l in combination with 75 mg/l of AgNO3. Embryogenesis and the regeneration rates reached 26.01 and 22.22%, respectively. The results indicated that the doubled haploid (DH) lines Jenah Khotifa and Biskri showed greater germination percentage with 70% compared to their parent with 60%. However, DH lines derived from crosses of Rezzak and Karim genotypes hybrid with maize had the same behaviour as their parent respectively. Also, the results showed that the DH lines equalled to the control for average height of plants and spikeless tillers/plant. The difference in average days from sowing to 50% ear emergence between DH1 lines from Biskri and Rezzak and the control was reduced to 5 and 9 days DHs lines showed 1000-kernel weight and spike tillers/plant greater in DH1 lines than in the control. However, kernels/spike was significantly greater in control compared with DH lines.
