Plant Virology Protocols: From Virus Isolation to Transgenic Resistance / Edition 1

Plant Virology Protocols: From Virus Isolation to Transgenic Resistance / Edition 1

by Gary D. Foster
Pub. Date:
Springer-Verlag New York, LLC

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Plant Virology Protocols: From Virus Isolation to Transgenic Resistance / Edition 1

The aim of Plant Virology Protocols is to provide a source of infor- tion to guide the reader through the wide range of methods involved in gen- ating transgenic plants that are resistant to plant viruses. To this end, we have commissioned a wide-ranging list of chapters that will cover the methods required for: plant virus isolation; RNA extraction; cloning coat p- tein genes; introduction of the coat protein gene into the plant genome; and testing transgenic plants for resistance. The book then moves on to treatments of the mechanisms of resistance, the problems encountered with field testing, and key ethical issues surrounding transgenic technology. Although Plant Virology Protocols deals with the cloning and expression of the coat protein gene, the techniques described can be equally applied to other viral genes and nucleotide sequences, many of which have also been shown to afford protection when introduced into plants. The coat protein has, however, been the most widely applied, and as such has been selected to illustrate the techniques involved. Plant Virology Protocols has been divided into six major sections, c- taining 55 chapters in total.

Product Details

ISBN-13: 9780896033856
Publisher: Springer-Verlag New York, LLC
Publication date: 02/24/1998
Series: Methods in Molecular Biology Series , #81
Edition description: 1998
Pages: 588
Product dimensions: 6.10(w) x 9.25(h) x 0.05(d)

Table of Contents

Part I. Introduction

Introduction to Plant Virology

Introduction to Classical Crossprotection

History of Coat Protein-Mediated Protection

Part II. Virus Isolation, RNA Extraction, and Location of Coat Protein Genes

Geminivirus Isolation and DNA Extraction

Caulimovirus Isolation and DNA Extraction

Reovirus Isolation and RNA Extraction

Procedures for Plant Rhabdovirus Purification, Polyribosome Isolation, and Replicase Extraction

Hordeivirus Isolation and RNA Extraction

Furovirus Isolation and RNA Extraction

Tobravirus Isolation and RNA Extraction

Tobamo virus Isolation and RNA Extraction

Potexvirus Isolation and RNA Extraction

Carlavirus Isolation and RNA Extraction

Potyvirus Isolation and RNA Extraction

Trichovirus Isolation and RNA Extraction

Ilarvirus Isolation and RNA Extraction

Bromovirus Isolation and RNA Extraction

Cucumovirus Isolation and RNA Extraction

Nepovirus Isolation and RNA Extraction

Comovirus Isolation and RNA Extraction

Carmovirus Isolation and RNA Extraction

Tymovirus Isolation and Genomic RNA Extraction

Tombusvirus Isolation and RNA Extraction

Luteovirus Isolation and RNA Extraction

Part III. Cloning of Coat Protein Genes

RNA Analysis: Size and 3' End Group Determination

RNA Fractionation by Density Gradient Centrifugation

cDNA Library Construction for Lambda ZAP®-Based Vectors

PCR Cloning of Coat Protein Genes

Antibody Production

Expression Library Screening

In Vitro Transcription and Translation

Analysis of Coat Expression Cassettes in Protoplasts

DNA Sequencing

Computer Analysis of Amino Acid Sequences: The Case of Plant Virus Capsid Proteins

Part IV. Plant Transformation and Primary Analysis

Preparation of Coat Protein-Containing Binary Vectors for Use in Agrobacterium-MediatedTransformation

Potato Transformation. Transformation of Tomato

Tobacco Transformation

Genetic Transformation of Wheat. Production of Transgenic Rice

Molecular Analysis of Transgenic Rice

PCR Analysis of Transgenic Tobacco Plants

Southern Analysis of Transgenic Tobacco Plants

Detection and Quantification of Transcript RNA in Transgenic Plants Using Digoxigenin-Labeled cDNA Probes

Western Analysis of Transgenic Plants

Part V. Evaluation of Resistance

Assaying Levels of Plant Virus by ELISA

Detection of Plant RNA Viruses by Nonisotopic Dot-Blot Hybridization

Detection and Quantification of Plant Viruses by PCR

Assaying Levels of Virus with Local Lesion Hosts

Field Testing Resistance of Transgenic Plants

Agronomic Performance of Transgenic Plants

Part VI. Mechanisms of Resistance

Mechanisms of Resistance: Expression of Coat Protein. Mechanisms of RNA-Mediated Resistance to Plant Viruses

Detection of Risks Associated with Coat Protein Transgenics

Potential Benefits of the Transgenic Control of Plant Viruses in the United Kingdom


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