A highly efficient transient protoplast system for analyzing defence gene expression and protein–protein interactions in rice

Molecular Plant Pathology - Tập 7 Số 5 - Trang 417-427 - 2006
Songbiao Chen1, LIZEN TAO2,1, Lirong Zeng1, Miguel E. Vega‐Sánchez1, Kenji Umemura3, Guo‐Liang Wang1
1Department of Plant Pathology, The Ohio State University, Columbus, Ohio 43210, USA
2College of Life Sciences, South China Agricultural University, Guangzhou 510642, China
3Meiji Seika Kaisha Ltd, Health & Bioscience Laboratories, 350-0289 Japan

Tóm tắt

SUMMARYThe transient assay system based on mesophyll or cultured cell‐derived protoplasts has been exploited in several plant species and has become a powerful tool for rapid gene functional analysis and biochemical manipulations. However, the system has not been widely used in rice owing to the difficulties in large‐scale isolation of viable rice protoplasts from leaves or suspension‐cultured cells. Here, we describe a significantly improved method to isolate a large number of protoplasts from stem and sheath tissues of both young and mature plants. High‐level coexpression of multiple constructs and efficient suppression of exogenous and endogenous genes were observed in the stem‐ and sheath‐derived protoplasts. A transient green fluorescent protein and luciferase‐based reporter system for defence‐related genes expression analysis has been established, which is useful for screening and characterizing genes involved in rice defence signalling pathways. Furthermore, a protoplast‐based bimolecular fluorescence complementation (BiFC) system for the detection of protein–protein interactions in living rice cells was developed. The YFP complementation of two split‐YFP halves mediated by homodimerization of the GUS and SPIN1, a cell‐death related protein, was observed in transfected protoplasts. In combination with genetic, genomic and proteomic approaches, the established versatile protoplast transient assay system will facilitate large‐scale functional analysis of defence‐related genes in rice.

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Tài liệu tham khảo

10.1111/j.1365-313X.1994.00421.x

10.1023/A:1024890601888

Asai T., 2000, Fumonisin B1‐induced cell death in Arabidopsis protoplasts requires jasmonate‐, ethylene‐, and salicylate‐dependent signaling pathways, Plant Cell, 12, 1823

10.1038/415977a

10.1111/j.1365-313X.2004.02206.x

10.1073/pnas.86.24.9692

10.1016/j.pbi.2005.09.011

10.1094/MPMI-18-0511

10.1007/BF01969712

10.1038/nbt0890-736

10.1111/j.1365-313X.2005.02586.x

10.1105/tpc.2.7.591

10.1094/MPMI.2004.17.2.140

10.1046/j.1365-313X.2002.01360.x

10.1023/B:PLAN.0000036368.74758.66

10.1046/j.1365-313X.2002.01291.x

10.1016/S1097-2765(02)00496-3

10.1038/nature03895

10.1105/tpc.105.037069

10.1104/pp.104.055624

10.1002/j.1460-2075.1987.tb02730.x

10.1007/BF00380613

10.1016/S0168-9452(96)04541-4

10.1073/pnas.97.6.2940

10.1038/27240

10.1038/ng1704

10.1007/s00299-002-0435-2

10.1046/j.1365-313X.2002.01394.x

10.1073/pnas.0404041101

10.1046/j.1365-313X.2003.01639.x

McElroy D., 1990, lsolation of an efficient actin promoter for use in rice transformation, Plant Cell, 2, 163

10.1093/oxfordjournals.pcp.a028918

10.1093/pcp/pch048

10.1093/nar/gnh180

Qu S. Liu G. Zhou B. Bellizzi M. Zeng L. Dai L. Han B.andWang G.L.(2006)The broad‐spectrum blast resistance gene Pi9 encodes an NBS‐LRR protein and is a member of a multigene family in rice.Genetics 172 1901–1914.

10.1046/j.0960-7412.2000.00942.x

Sambrook J., 2001, Molecular Cloning: a Laboratory Manual.

10.1016/j.febslet.2005.06.077

10.1104/pp.010820

10.1038/338274a0

10.1046/j.1365-313X.1999.00405.x

10.1105/tpc.006320

10.1111/j.1365-313X.2004.02219.x

10.1094/MPMI.2000.13.8.869

10.1105/tpc.104.025171

10.1093/nar/gkj016

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Thông tin xuất bản

Molecular Plant Pathology

Tập 7 Số 5

417-427

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