Engineering of rice varieties with enhanced resistances to both blast and bacterial blight diseases via CRISPR/Cas9.

Rice blast and bacterial blight represent two of major diseases having devastating impact on the yield of rice in most rice-growing countries. Developments of resistant cultivars are the most economic and effective strategy to control these diseases. Here, we used CRISPR/Cas9-mediated gene editing to rapidly install mutations in three known broad-spectrum blast-resistant genes, Bsr-d1, Pi21 and ERF922, in an indica thermosensitive genic male sterile (TGMS) rice line Longke638S (LK638S). We obtained transgene-free homozygous single or triple mutants in T1 generations. While all single and triple mutants showed increased resistance to rice blast compared with wild type, the erf922 mutants displayed the strongest blast resistance similar with triple mutants. Surprisingly, we found that Pi21 or ERF922 single mutants conferred enhanced resistance to most of tested bacterial blight. Both resistances in mutants were attribute to the up-regulation of SA- and JA-pathway associated genes. Moreover, phenotypic analysis of these single mutants in paddy fields revealed that there were no trade-offs between resistances and main agricultural traits. Together, our study provides a rapid and effective way to generate rice varieties with resistance to both rice blast and bacterial blight.

A Procedure for Inducing the Occurrence of Rice Seedling Blast in Paddy Field.

Rice blast caused by the filamentous fungus Magnaporthe oryzae, is arguably the most devastating rice disease worldwide. Development of a high-throughput and reliable field blast resistance evaluation system is essential for resistant germplasm screening, resistance genes identification and resistant varieties breeding. However, the occurrence of rice blast in paddy field is easily affected by various factors, particularly lack of sufficient inoculum, which always leads to the nonuniform occurrence and reduced disease severity. Here, we described a procedure for adequately inducing the occurrence of rice seedling blast in paddy field, which involves pretreatment of diseased straw, initiation of seedling blast for the first batch of spreader population, inducing the occurrence of the second batch of spreader population and test materials. This procedure enables uniform and consistent infection, which facilitates efficient and accurate assessment of seedling blast resistance for diverse rice materials.