| Summary: | 碩士 === 國立中興大學 === 生物科技學研究所 === 101 === Bamboo mosaic virus (BaMV) is a single-stranded positive sense RNA virus. To study the infection cycle of BaMV in its assay host, Nicotiana benthamiana, we have used cDNA-AFLP technique to identify the differentially expressed genes after BaMV infection. The first project of my thesis is analyzing one downregulated gene, ACAC3, and two upregulated genes, ACAC9-1 and ACCT6-1, to examine whether these genes are involved in the infection of BaMV in N. benthamiana plants. I have used virus induced gene silencing technique to knock down the expression levels of these three genes, and then inoculated BaMV on the knockdown N. benthamiana leaves. The accumulation levels of virus coat protein are measured by Western blotting analysis. The results showed that the accumulation of BaMV was increased to 113% in ACAC3 knockdown plants at 5 dpi compared to that of the control plants. However, the accumulation levels of BaMV coat protein in ACAC9-1 (135%) and ACCT6-1 (60%) knockdown plants were increased and decreased, respectively, significantly to that of control plants at 5 dpi. The results imply that the two upregulated gene ACAC9-1 and ACCT6-1 may play different roles as restricting and assisting, respectively, in BaMV infection cycle. The second project of my thesis is characterizing an up-regulated gene ACCT8-1 which was studied previously. This gene was involved in the movement of BaMV in N. benthamiana plants. The identity of ACCT8-1 is annotated to an elicitor inducible leucine-rich repeats receptor-like protein of N. tabacum and the tomato homologous of cladosporium fulvum resistance gene Cf-2 (Hcr2). To study the interaction of ACCT8-1 with BaMV in the infection cycle, I generated a deletion construct of ACCT8-1 which removed the leucine-rich repeats domain (ΔLRR). I transiently expressed ACCT8-1/ΔLRR-GFP in BaMV inoculated N. benthamiana. The accumulation of BaMV coat protein was enhanced in ACCT8-1/ΔLRR-GFP expressed N. benthamiana at 5 dpi. The results indicate that the leucine-rich repeats domain is not essential for BaMV movement.
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