Generation and application of monoclonal antibody against Phalaenopsis chlorotic spot virus and polyclonal antibody against Carnation mottle virus Phalaenopsis orchid isolate

• Main Authors: Cheng-quan Chen, 陳呈權
• Other Authors: Fuh-Jyh Jan
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/45620180441199567091

碩士 === 國立中興大學 === 植物病理學系所 === 97 === Abstract In 2003, an isolate of the potyvirus Phalaenopsis chlorotic spot virus (PhCSV) and a phalaenopsis isolate of the carmovirus Carnation mottle virus (CarMV-Ph) were isolated from Phalaenopsis orchid plants. On the infected plants, these virus isolates caused respectively leaf chlorotic spot and leaf chlorotic ring symptoms. Despite reported presence of these viruses in Phalaenopsis orchid for several years, these accurate detection from the infected plants was ineffective, because of lack of specific and sensitive detection tools. Hence, to develop an officient a detection tool, antibodies against PhCSV and CarMV-Ph particles were generated. Purified PhCSV was injected into BALB/c mice for the production of hybridomas from mice spleen cells with FOX-NY myeloma cells by PEG-1450, and the resultant positive hybridoma clones were screened by indirect ELISA. Three cell lines were selected and determined to be IGg1/κ isotype. The titer of the monoclonal antibody (106) and poltclonal antibody (105) was determined by indirect ELISA, and the specificity of the antibody to the antigen was determined by western blotting analysis. Polyclonal antibody against the CarMV-Ph was produced in a New Zealand white rabbit. After 4 rounds of immunization by purified CarMV-Ph virus particles, the rabbit was bled weekly and antiserum was collected. The generated PhCSV monoclonal antibody and CarMV-Ph polyclonal antibody have effective application potentials in the immunological detection and can be employed to detect these economically important viruses in Phalaenopsis orchid nursery to produce virus-free Phlalenopsis orchid seedlings.