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A Study on Defense Mechanism of Maize Seedlings to Drought Stress Mediated by microRNA

Author: WeiLiYa
Tutor: ZhangZuXin
School: Agricultural University of Hebei
Course: Crop Genetics and Breeding
Keywords: Zea mays drought stress miRNA antioxidant defense stomatal closure osmoprotection
CLC: S513
Type: Master's thesis
Year: 2009
Downloads: 256
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Abstract


Drought is one of environmental factors that greatly impact plant distribution as well as crop production. An effective approach to increase the crop production is to utilize drought-tolerant crops. Using molecular genetics and genetic engineering to obtain the drought-tolerant crops has been one of the most important goals in modern agriculture. Therefore, understand of the molecular basis of plant response to drought stress will be greatly important for genetically improving the drought tolerance of crops. In the long course of evolution, plants express a lot of key genes involved in defense mechanism to resist drought stress. Under drought condition, not only the protein-coding genes, but also the non-protein-coding genes (such as microRNA: miRNA) were induced for response. In order to resist drought stress, expression of miRNA (miRNA) genes will be induced and their transcripts (miRNAs) can regulate gene expression by guiding target mRNA cleavage or translation inhibition. Finally, miRNAs participates in initiate the morphological and physiological mechanisms of drought stress tolerance.A customμParaflo? microfluidic chip containing 623 unique plant probes from version 10.0 was used in maize (Zea mays) inbred line, X178, which is a drought-tolerant elite inbred line in china, was used in this study. We searched drought stress responsive miRNAs in maize leaves, and analyzed the expression profile of miRNAs and target genes under stress condition. Our results are mainly as follows:1. A total of 34 miRNAs, representing 5.46% (34/623) of the probes on the microarray in maize leaves under drought condition, were changed at the 1% significance level. Eleven out of 34 differentially expressed miRNAs were aligned with eight maize miRNA families, while the other miRNAs corresponded to members of eleven miRNA families from another six plant species.2. The clustering results indicated that 34 drought responsive miRNAs had been repressed or induced, and three predominant expression patterns were detected at different time points during the drought stress process. The expressions of 8 miRNAs were up-regulated and recovered to normal levels during late treatment. However, 10 miRNAs were down-regulated and then up-regulated. In addition, sixteen miRNAs were repressed after drought treatment. These results strongly suggested that different response of miRNAs might contribute to the regulation at different stages during the drought process.3. Based on bioinformatics method 32 target genes of drought-responsive miRNAs were identified and given functional annotations. Biological functions of the predicted target genes were classified into two categories, functional proteins and regulatory proteins. Motifs in the 5′-upstream region of 32 predicted target genes were queried by the PlantCARE motif-finding algorithm. The most frequently (65.6%) appearing cis-acting element was the ABRE, which was present in the 5′- upstream region of 21 out of 32 targets, indicating that these miRNA-mediated genes are involved in the regulation of the transduction of stress signals in drought-induced ABA cascade. Of the predicted targets, the expression profiles of eight genes were confirmed by RT-PCR. The expression profiles of the eight target genes showed a consistent with their corresponding miRNAs function.4. We detected the content of ABA and free proline, POD activity, and the expression profile of miR474-mediated proline dehydrogenase gene (PDH), miR528-mediated peroxidase gene (POD), miR168a-mediated mitogen-activated protein kinase gene (MAPK) and miR167d-mediated phospholipase D gene (PLD). Combining the expression of miRNAs and target genes with the activity of proteins which target gene code, we suggested that miRNAs potentially participate in the post-transcriptional regulation of genes responsive to drought stress in maize seedlings. It was indicated that a hypothesis on defense mechanism of maize seedlings to drought stress mediated by miRNA.

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