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为了深入研究作物耐逆境胁迫的分子机制和发现新的水稻耐逆基因,采用Affymetrix基因芯片技术与水稻表达芯片(含51,279个转录本),分析了培矮64S不同生长发育时期、不同组织器官全基因组在低温、干旱、高温逆境胁迫下的表达水平变化,筛选出一批耐多逆境候选功能基因。OsMsr13是其中一个受低温、高温与干旱诱导,在各生长发育时期与组织器官均显著上调的基因。根据序列生物信息学分析,OsMsr13含有7个外显子和6个内含子c,DNA全长序列为1603bp,完整的ORF为1317bp,编码一个含438个氨基酸残基的蛋白质;在OsMsr13启动子区域发现多个与逆境应答相关的顺式作用元件;数据库查找和蛋白质多序列的比对表明OsMsr13的预测蛋白属于钙调素结合蛋白家族。为了进一步研究OsMsr13基因的功能,通过RT-PCR方法扩增到包含OsMsr13完整ORF的cDNA克隆,构建了含有OsMsr13重组基因的过量表达载体pCAM-JIT-OsMsr13,并经农杆菌介导法将重组基因导入到水稻中。转OsMsr13株系苗期的抗逆性试验表明OsMsr13过量表达能显著地提高水稻的耐寒性,但在抗旱和耐高温方面,转基因植株和野生型9311没有观察到多大差别。
In order to further study the molecular mechanism of crop stress tolerance and find new rice stress tolerance genes, Affymetrix microarray and rice expression chips (including 51,279 transcripts) were used to analyze the effects of different growth and development stages of Pei’ai 64S The genome was changed under the stress of low temperature, drought and high temperature stress, and a number of functional candidate genes for resistance to drought stress were screened out. OsMsr13 is one of the genes that is induced by low temperature, high temperature and drought and significantly up-regulated in all tissues and organs during the growth and development. According to the sequence bioinformatics analysis, OsMsr13 contains 7 exons and 6 introns c, the full-length DNA sequence is 1603bp, the complete ORF is 1317bp, encodes a protein with 438 amino acid residues; OsMsr13 promoter A number of cis-acting elements were found in the region that were associated with adversity responses; the alignment of the database search with the protein multiple sequence indicated that the predicted protein of OsMsr13 belongs to the calmodulin binding protein family. In order to further study the function of OsMsr13 gene, a cDNA clone containing the complete ORF of OsMsr13 was amplified by RT-PCR. The overexpression vector pCAM-JIT-OsMsr13 containing OsMsr13 recombinant gene was constructed and transformed into E.coli BL21 Imported into rice. The resistance test of transgenic OsMsr13 at seedling stage showed that overexpression of OsMsr13 could significantly increase the cold tolerance of rice, but no significant difference was observed in transgenic plants and wild type 9311 in terms of drought resistance and high temperature tolerance.