Expression of Lily mottle virus Coat Protein and Preparation of IgY Antibody against the Recombinant Coat Protein

• Published in: Weon'ye gwahag gi'sulji Vol. 32; no. 4; pp. 544 - 549
• Main Authors: Yoo, Ha Na, Jung, Yong-Tae
• Format: Journal Article
• Language: English
• Published: 한국원예학회HST 01.08.2014; 한국원예학회
• Subjects: 농학; IC-RT-PCR; egg yolk immunoglobulin
• ISSN: 1226-8763; 2465-8588
• DOI: 10.7235/hort.2014.13167

Lily symptomless virus (LSV), Lily mottle virus (LMoV), and Cucumber mosaic virus (CMV) are the most prevalent virusesinfecting lilies in Korea. Leaf and bulb samples showing characteristic symptoms of virus infection were collected in 2012, and80 field samples were analyzed by reverse transcription polymerase chain reaction (RT-PCR). The infection frequencies were79% for LMoV, 5% for LSV, and 3% for CMV. The LMoV coat protein gene was amplified and cloned into the pET21d(+) expressionvector to develop serological diagnostic tools to detect LMoV. The resulting carboxy-terminal His-tagged coat proteins were expressedin Escherichia coli strain BL21 (DE3) by induction with IPTG. The recombinant proteins were purified using Ni-NTA agarosebeads and used as an antigen to produce polyclonal antibodies in laying hens. The resulting egg yolk immunoglobulin (IgY)specifically recognized LMoV from infected plant tissues in immunoblotting assays and had comparable sensitivity to that ofa mammalian antibody. In addition, method of immunocapture RT-PCR using this IgY was developed for sensitive, efficient,and rapid detection of LMoV. Based on these results, large-scale bulb tests and detection of LMoV in epidemiological studiescan be performed routinely using this IgY. This is the first report of production of a polyclonal IgY against a plant virus andits use for diagnosis. KCI Citation Count: 2