The study of the ACTG7-1 from Nicotiana benthamiana involving in the replication of Bamboo mosaic virus

• Main Authors: Hsiao-Chen Wang, 王筱禎
• Other Authors: Ching-Hsiu Tsai
• Format: Others
• Language: en_US
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/86063961401658588098

碩士 === 國立中興大學 === 生物科技學研究所 === 100 === Bamboo mosaic virus (BaMV) is a single-stranded positive sense RNA virus. It belongs to Potexvirus of Flexiviridae. Since the virus encodes a limit number of genes can not complete its infection cycle by itself, it requires host factors at different stages of the infection, such as translation, replication, and movement. Previously, our lab used cDNA-amplified fragment length polymorphism (cDNA-AFLP) technique to identify the differentially expressed gene fragments from BaMV infected Nicotiana benthamiana plants. One of the downregulated cDNA fragments, ACTG7-1, is an ortholog to REVERSION-TO-ETHYLENE SENSITIVITY1 (RTE1) of Arabidopsis. According to the previous study, the expression of ACTG7-1 was knocked down by virus-induced gene silencing technique; the coat protein accumulation level of BaMV was increased about 4 folds to that of control plant at 5 days of post inoculation. To examine whether ACTG7-1 is involved in replication step of the BaMV infection cycle, I prepared the ACTG7-1 knockdown protoplasts derived from knockdown plants and inoculated with BaMV viral RNA. Results indicates that the coat protein accumulation is increased to 2-3 folds that of the control protoplasts suggesting a negative role of ACTG7-1 in BaMV replication. Further analysis of the accumulation of BaMV plus- and minus-strand genomic RNAs by real-time RT-PCR suggests the effect on minus-strand RNA synthesis. Transiently expressed the fusion protein of ACTG7-1 with orange in N. benthamiana shows the ER localization by confocal microscopy. To test whether ACTG7-1 affected the replication of BaMV through the ethylene pathway, we have used the ethylene precursor ACC to enhance the ethylene response. The result indicated ethylene might enhance the resistance against BaMV. The relationship of BaMV replication and ACTG-7-1 in N. benthamiana is still needed to be clarified.