Development of a High-Throughput Screening Method Using a Cell-Based, Lawn Format Assay for the Identification of Novel Plant Defense Activators from Combinatorial Peptide Libraries

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• 作者：Masahiro Miyashita ; Yukari Otake ; Masashi Oda ; and ; Hisashi Miyagawa
• 刊名：Journal of Agricultural and Food Chemistry
• 出版年：2007
• 出版时间：February 7, 2007
• 年：2007
• 卷：55
• 期：3
• 页码：806 - 811
• 全文大小：286K
• 年卷期：v.55,no.3(February 7, 2007)
• ISSN：1520-5118

文摘

Plants respond to attack by pathogens through various defense mechanisms. These defenseresponses are triggered by a variety of molecules derived from pathogenic microorganisms as wellas host plants. In this study, we developed a high-throughput screening method using a cell-basedlawn format assay for the identification of novel peptides that can induce plant defense responsesfrom combinatorial peptide libraries. Solid-phase peptide libraries were synthesized using aphotocleavable linker and immobilized using agarose gel. The peptides were partially cleaved frombeads, and the agarose gel was layered on the tobacco cells. The defense response was thenobserved by detecting the generated H₂O₂ using a sensitive H₂O₂ indicator dye, N-(carboxymethylaminocarbonyl)-4,4'-bis(dimethylamino)diphenylamine sodium salt (DA-64). Using this assay format,a 6859-member peptide library based on the sequence of flagellin-derived peptides was screened,and several structural features important for the activity were obtained.