Research in Plant Disease 2012;18(3):231-235.
Published online September 30, 2012.
Indian citrus ringspot virus의 ELISA 진단 시스템 구축
신명주 ( Myeung Ju Shin ) , 권영철 ( Young Chul Kwon ) , 노현수 ( Hyeon Su Ro ) , 이현숙 ( Hyun Sook Lee )
Construction of ELISA System for the Detection of Indian citrus ringspot virus
Myeung Ju Shin, Young Chul Kwon, Hyeon-Su Ro and Hyun-Sook Lee*
Department of Microbiology and Research Institute for Life Science, Gyeongsang National University, Jinju 660-701, Korea
Received: March 12, 2012   Revised: June 26, 2012   Accepted: June 30, 2012
Indian citrus ring spot virus (ICRSV) is known to cause a serious disease to citrus, especially to Kinnow mandarin, the popular cultivated citrus species in India. In this study, we developed diagnostic systems based on enzyme-linked immunosorbent assay (ELISA). In order to generate antibodies against ICRSV coat protein, we overexpressed the coat protein in Escherichia coli using the pET15b expression vector containing an optimized ICRSV coat protein gene. The recombinant ICRSV coat protein was overexpressed as soluble form at 37oC upon IPTG induction. The protein was purified to 95% in purity by Ni-NTA column chromatography. The purified protein was immunized to rabbit for the generation of polyclonal antibody (PAb). The PAb showed a specific immunoreaction to recombinant ICRSV coat protein in western blot analysis and ELISA. Diluted rabbit antisera (10,000 fold) could detect less than 10 ng and 5 ng of the target protein in western blot and ELISA analysis, respectively.
Key Words: Diagnosis, ELISA, Indian citrus ringspot virus, Polyclonal antibody
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