MiR444b.2 And RPW8.1 Regulate The Disease Resistance To Rice Blast And The Development Of Rice Growth

MicroRNA is a class of non-coding small molecular RNA,which length is 20-24 nt.It was widespread in eukaryotes.In plants,microRNA may be involved in the physiological processes such as growth and development,biotic stress and abiotic stress.In our previous study,through the comparative analysis of accumulation of microRNA in susceptible(LTH)and resistant(IRBLkm-Ts)rice accessions was significantly different in response to Magnaporthe oryzae infection,a series of microRNA was screened that may be involved in rice immunity to agaist Magnaporthe oryzae.Deep sequencing data showed that,after treating with rice blast fungus,with the passage of time,the accumulation of miR444b.2 in LTH was increased gradually,but decreased in IRBLkm-Ts.We constructed miR444b.2 over-expression transgenic line and identification of resistance to rice blast disease as well as its agronomic traits.The results showed that:first,after inoculation with Magnaporthe oryzae,over-expression of miR444b.2 transgenic rice plants were more susceptible rice blast disease.It was showed that there was more larger and much number lesions in over-expression of miR444b.2 transgenic lines leaf than in control rice plant;Expression of defense related genes were significantly down-regulated in miR444b.2 transgenic rice lines compared with control lines;The accumulation of H2O2 was less or no in miR444b.2 transgenic rice lines than in control rice lines.Second,during cultivating in the field,a spontaneous event happens because that was infected rice sheath blight disease on the leaf sheath surface of homozygous T3 plants in miR444b.2 transgenic accessions.The data showed that there were 62%and 53%plants susceptible rice sheath blight disease on 1#and 3#lines respect;there was 20%plants susceptible rice sheath blight disease on contorl line.For the further study,On the other hand,after analysis of the rice sheath blight disease rating,it was significant higher in transgenic line than in control line.In the third,after statistical analysis of the rice tiller,the transgenic rice plants tillers of over-expression of miR444b.2 were significantly decreased compared with the control group.The results showed that miR444b.2 is involved in negative regulation of rice blast and sheath blight disease resistance,and negative regulation of rice tiller number.Molecular mechanism of rice blast disease resistance has been a research hot spot,the identification of rice blast resistance genes mainly belong to encode NBS-LRR protein gene,according to the gene for gene hypothesis,this kind of resistance protein has many small kind of specific resistance,and therefore resistance do not endure,therefore,it is necessary to excavate broad-spectrum resistance genes.RPW8.1 is one of a gene which was located in RPW8 loci in Arabidopsis thaliana.And there was stronger disease resistance to powdery mildew and peronospora parasitica?belong to a special kind of broad-spectrum resistance genes.So,We used the induction promoter PR10 to express RPW8.1 after inoculating rice blast fungus in rice,by constructing heterologous expression of RPW8.1 transgenic rice plants to confirm the RPW8.1 transgenic rice disease resistance to Magnaporthe oryzae and analysis the agronomic traits.The study found that after inoculating with M.oryzae,RPW8.1 transgenic rice plants showed stronger disease resistance to Magnaporthe oryzae;during cultivating in the field,the heterologous expression of RPW8.1 homozygous rice plants were significant difference in agronomic traits compared with the control group.It showed that:First,after inoculation of Magnaporthe oryzae,the lesion area of leaves of RPW8.1 transgenic rice was significantly reduced compared with the control rice plants,the grade of rice blast disease in leaf was lower than the control plants;the defense related genes was significantly up-expressed compared with control rice lines;Second,after treatment with Magnaporthe oryzae and chitin elicitor,RPW8.1 and defense related genes significantly induced in RPW8.1 transgenic rice lines;Third,after heterologous expression of RPW8.1 in rice plants,there was lower plant height,shorter spike length,weaker seed setting rate,lighter 1000-grain weight,shorter grain length and smaller grain width compared with control rice plants.The results showed that RPW8.1 gene can significantly enhance the rice disease resistance to rice blast fungus;As the same time,the effect of the agronomic character in rice was significant.