| This thesis includes two parts: 1)The study on the function of OsWRKY97 gene in Rice.2)clone and functional analysis of OsDUF946.4.Rice is one of the important crops,however,with the continuous deterioration of the environment,drought,salinity,temperature are important factors to limit the growth of plants,but also the main abiotic stress factors affecting crop yield,so it is particularly important to study the mechanism of plant response to abiotic stress.At present,the focus of rice genome research has shifted from structural genomics to functional analysis.The discovery of new genes and the annotation of gene functions are considered to be an important issue in the study of functional genomics.In this study,we explored the function of two rice genes.The main research contents and results are as follows:1.WRKY transcription factor has been found to play an important role in rice biological development,biological stress and abiotic stress.Here,we identified a new member of WRKY transcription factor family,OsWRKY97,and analyzed its role in abiotic stress.It was found that the drought tolerance of OsWRKY97 overexpressed plants was higher than that of the wild-type(WT)plants at the germination,post-germination and seedling growth stages.These results suggest that OsWRKY97 was positively regulates drought tolerance in rice.The transient expression of OsWRKY97-GFP fusion protein driven by 35 S promoter in tobacco cells indicated that OsWRKY97 was located in the nucleus.Our study also found that OsWRKY97 expression plants also showed higher sensitivity to exogenous abscisic acid(ABA)at germination and post-germination stage compared with the WT.Therefore,we speculated that OsWRKY97 regulate the drought tolerance of rice through ABA dependent pathway.The results showed that the overexpression of OsWRKY97 increased the accumulation of ABA and slowed down the rate of water loss.This result confirms the above conjecture.In addition,we also found that the content of proline in OsWRKY97 overexpressed plants was higher than that in the WT,and the levels of malondialdehyde(MDA)and reactive oxygen species(ROS)were lower.Active oxygen scavenging enzymes including superoxide dismutase(SOD),catalase(CAT)and peroxidase(POD)increased to maintain the homeostasis of ROS in cells.All these results indicate that OsWRKY97 plays an important role in rice response to drought stress and has potential application value in improving rice drought resistance.2.Domain of unknown function(DUF)is a kind of protein domain,most of which are conservative.Some unique DUFs in plants are involved in biological processes such as plant growth and development,defense response to pests and diseases,and response to abiotic stress.In this study,we cloned a adversity candidate gene OsDUF946.4,which has an open reading frame of 1653 bp and encodes 550 amino acid.The molecular weight of the peptide encoded by OsDUF946.4 is 59 k Da,and its isoelectric point is 6.1.Based on the analysis of online promoters of Plant CARE,it is found that the promoters of OsDUF946.4 have many homeostasis elements related to adversity.The expression pattern of OsDUF946.4 gene in rice was analyzed by quantitative q RT-PCR.It was found that the expression of OsDUF946.4gene can be induced by different adversity,which belongs to multi adversity induced gene.In order to further verify its role in abiotic stress,we constructed p CAMBIA1301S-OsDUF946.4 overexpression vector,p CAMBIA1305-OsDUF946.4 promoter expression vector,p CAMBIA2300-e GFPOsDUF946.4 overexpression vector and p ET32a(+)-OsDUF946.4 expression vector.And then OsDUF946.4-overexpression plants and promoter plants were obtained.Through the transient expression of Agrobacterium mediated tobacco cells,OsDUF946.4 subcellular was located in plasmalemma and nucleus.In addition,we also found that the expression of pET32a-OsDUF946.4 fusion protein in E.coli can increase its drought tolerance and salt tolerance.These results preliminarily confirm that OsDUF946.4 play an important role in the response of abiotic stress. |
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