Molecular Marker-assisted Selection Breeding Using The Broad-spectrum Blast Resistance Gene Pi9in Rice

Rice blast, caused by the fungal pathogen Magnaporthe oryzae, is one of the most important diseases in rice production. Development and application of resistant rice varieties harboring broad-spectrum blast resistance gene(s) is the most effective, economical and environment-friendly strategy to control the damage of this disease. To improve blast resistance of twenty receptor indica rice lines, polymorphic DNA markers were developed based on the sequence of the cloned broad-spectrum blast resistance gene Pi9and marker-assisted selection breeding practice was implemented in this study. The main results were as follows:1. Effects of three media on the growth of Magnaporthe oryzae was investigated, and oat-rice leaf medium and oat-tomato medium could accelerate the growth of mycelium, but the quality of mycelium cultured on three tested media showed no significant difference. On the other hand, the sporulation ability of different Magnaporthe oryzae races or strains on the same medium showed a big difference. Among fifty-one tested races or strains, thirty-five showed excellent sporulation ability, three showed poor sporulation ability, and thirteen couldn’t produce spores at all.2. Blast resistance investigation in field nursery showed that75-1-127, the donor of the Pi9gene, expressed high level resistance, while all receptors showed different grades of susceptibility. Greenhouse inoculation assay showed that75-1-127was resistant to83.3%of twenty-four Magnaporthe oryzae races or strains collected from different parts of the world, while the resistance ratios of twenty receptors ranged from25%to62.5%. These results implied that blast resistance of all tested receptor rice lines is needed to be improved.3. Five innergenic polymorphic dominant molecular markers, INS1-1, INS2-1, INS1-3, INS2-3and pi9utr, were developed based on the genomic DNA sequence of the Pi9gene. The polymorphism ratios tested by INS2-1and INS2-3between two parents of twenty crosses are100%, and polymorphism ratios tested by INS1-1, INS1-3and pi9utr are75%.4. The genotype-assisted selection efficiency for blast resistance phenotype was over 95%by any of above four markers, INS1-1, INS2-1, INS2-3and pi9utr, resulting from randomly sampled individuals analyses of different backcross populations. The Pi9gene showed high level expression in all Fi9-gene present and resistant plants, and Pi9gene expression couldn’t be checked in all Pi9-gene absent and susceptible individuals, according to randomly sampled individuals analyses of two backcross populations,316B X75-1-127BC4F1and R288X75-1-127BC6F1.5. Above molecular markers were employed for marker-assisted selection in consecutive backcross breeding to improve blast resistance of the tested receptor rice lines, and one BC7F! population, six BC6F2populations, four BC6F1populations, four BC5F1populations and five BC4F1populations were developed, which paved a good foundation for further breeding elite blast resistant rice varieties.