Identification and functional characterizationof virus resistant genes of cucurbits by proteomics technologies

• Main Authors: Chang-Yang Liao, 廖昌洋
• Other Authors: Chung-Chi Hu
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/38675669967906740380

碩士 === 國立中興大學 === 生物科技學研究所 === 96 === Viral diseases are the major limiting factor for the production of cucurbits in Taiwan and worldwide. The use of resistant cultivars presents one of the most effective and economical approaches in viral disease management. For wax gourds, a cultivar TVI4204, originated from India has been shown to be immune to the challenge of Papaya ringspot virus watermelon type (PRSV-W) and Zucchini yellow mosaic virus (ZYMV) by Dr. Deng and Lin in Taiwan Agricultural Research Institute. The progeny of TVI4204 has also been shown to be highly resistant to most of the RNA viruses infecting cucurbits, and previous results suggested the resistance to PRSV-W is controlled by a single recessive gene, which is highly significant in molecular virology and practical applications. Therefore, the aim of this study is to identify the resistant gene through proteomics approaches, including two dimensional gel electrophoresis (2-DE) and mass spectrum analysis. A susceptible cultivar, TVI11577, with very similar genetic background, is used as a negative control. Accordingly, the resistant mechanisms involved in defense against the viral infection were studied further. The comparison of protein profiles obtained by 2-DE from TVI4204 and TVI11577 revealed changes in several proteins. From differentially expressed protein spots, the identifies of proteins were analyzed by LC MS/MS. Of these proteins, fructose-bisphosphate aldolase, glycine-rich RNA binding protein, and 14-3-3 family protein were induced more in TVI4204, whereas superoxide dismutase proteins were expressed more in TVI11577. The results indicate that the majority of the identified proteins are involved in regulate cellular metabolism, environmental stress, signal transduction and defense reactions. For those proteins without known functions, R2(pI：5.7；Mw：16 kD), R4(pI：6.9；Mw：18 kD), S1(pI：6.6；Mw：17 kD),S5(pI：5.9；Mw：12 kD), mR1(pI：5.3；Mw：34.5 kD), mR2(pI：5.1；Mw：27 kD) were identified to be homologous of Photosystem I reaction center subunit IV、Thioredoxin M－type、unnamed protein product、Os07g0150200、ORF1 of Cucumis sativus and Chlorophyll a-b binding protein of LHCII type I. In the further work, we want to find the host factor that can be binding with virus genome by using Oligo-dT cellulose column. Maybe can use geminivirus to development a viral vector and to use RNAi strategy to evaluate the function of the genes of interest.