Cloning And And Functional Study Of OsMYB105, A Node Gene Of Rice Drought Tolerence Regulatory Network

Rice is the main food crop in China,and more than half of the people take rice as a ration.Rice needs a lot of fresh water resources in the production process,and China is a country with water shortage.It is an important way to develop water-saving agriculture and ensure the safety of food production to discover drought-tolerant genes of rice,analyze its regulatory mechanism,and cultivate water-saving and drought-resistant rice.In this study,a gene Os MYB105(LOC_Os05g40960)in the co-expression network node of rice drought tolerance differentially expressed genes obtained by transcriptome analysis was used for gene cloning,transgenic rice cultivation and functional research.The main results are as follows:1.A drought-tolerant candidate gene,Os MYB105,was cloned from Zhenshan 97B(ZS97B).Os MYB105 encodes a MYB transcription factor(TF),whose genetic relationship is slightly near to the 1R type MYB transcription factor and far from other types of MYB TFs.There are differences in the sequence of this gene and between the indica rice cultivar ZS97B(Os MYB105ZS97)and the japonica rice cultivar NIP(Os MYB105NIP).2.The tissue expression pattern analysis showed that Os MYB105 was mainly expressed in young leaves.The promoter region of Os MYB105 contains ABRE,MBS and other response cis-elements related to abiotic stress.The quantitative PCR results showed that expression of Os MYB105 was induced by various abiotic stress(drought stress,cold stress,heat stress and PEG stress)and ABA treatment.3.The CRISPR/Cas9 gene knockout vector and overexpression vector of Os MYB105 were constructed,and the Os MYB105 gene overexpression and knockout transgenic rice lines were obtained.Overexpression of Os MYB105 can significantly increase the germination rate of rice seeds under PEG and NaCl stress,and reduce the sensitivity to ABA;knockout of Os MYB105 significantly reduces the germination rate of rice seeds under Na Cl and ABA treatments.Overexpression of Os MYB105 also enhanced tolerance to mannitol osmotic stress of transgenic rice plants.4.The drought tolerance of Os MYB105 transgenic plants was identified,and it was found that after drought treatment and rehydration,the survival rate of the overexpression lines was significantly higher than that of the wild-type control lines.Physiological tests showed that the proline content of the overexpressed lines was significantly higher compared with that of wild-type rice,and the water loss rate of leaves was reduced.On the contrary,the survival rate of the osmyb105 mutant under drought stress was significantly lower than that of the wild-type rice.The proline content of osmyb105 mutant was significantly lowed compared with that of wild-type rice plants,and the leaf water loss rate was increased.It indicated that Os MYB105 could positively regulate drought tolerance in rice.5.Subcellular localization analysis showed that Os MYB105 was located in the nucleus,Os MYB105 has transcriptional auto-activation activity in yeast,and can form homodimer.Transcriptome sequencing results showed that compared with wild-type rice,the expression of a number of genes related to abiotic stress,including the late embryogenesis abundant protein(LEA)-encoding gene,was up-regulated in Os MYB105 overexpressed transgenic rice.The promoters of some of these differentially expressed genes contained MYB binding sites,indicating that Os MYB105 may be participate in the response of rice to drought stress through regulating the expression of these genes.