Molecular plant biology / edited by Philip M. Gilmartin and Chris Bowler.

Format:

Book

Contributor:

Gilmartin, P. M.

Bowler, Chris, Dr.

Series:

Practical approach series ; 258.

Practical approach series ; no. 258-259.

The practical approach series ; no. 258-259

Language:

English

Subjects (All):

Plant molecular biology.

Physical Description:

volumes : illustrations (some color) ; 26 cm.

Place of Publication:

Oxford ; New York : Oxford University Press, 2002-

Contents:

Section 1 Gene Identification

1 Classical mutagenesis in higher plants / Maarten Koornneef 1

3 Choices in a mutagenesis experiment 4

Parental genotypes 4

Choice of mutagens 4

Mutagen dose 6

Numbers of M1 and M2 plants 8

The handling of M1 plants 9

4 Further analysis of mutants 9

2 Plant transformation / Karabi Datta, Swapan K. Datta 13

Embryogenic culture and recalcitrance 14

2 Agrobacterium-mediated transformation 15

Agrobacterium-tumefaciens and the Ti plasmid 15

Vector systems for Agrobacterium-mediated transformation 15

Supervirulent strains and superbinary vectors 16

Transformation using Agrobacterium as a vector 16

3 Transformation by biolistic method 20

Biolistic transformation 23

4 Protoplast transformation 25

Protoplast transformation using polyethylene glycol (PEG) 26

3 T-DNA tagging / Csaba Koncz, Jeff Schell 33

2 Generation of T-DNA transformants 34

The mechanism of T-DNA transformation 34

Agrobacterium hosts 34

3 T-DNA tagging in Arabidopsis 35

Generation of transformants 35

4 Screening approaches and theoretical considerations 43

Phenotype-based screening 43

Reverse genetics using sequence-based PCR screening approaches 45

Identification of gene mutations using sequenced T-DNA tags 46

4 Transposon tagging methods in heterologous plants / Andy Pereira 53

2 Generation of lines for transposon tagging 54

Phenotypic excision assays 54

Autonomous Ac systems 58

Two-Component Ac-Ds tagging systems 59

En-I(Spm) tagging systems 61

3 Mapping transposon lines 62

4 Transposon tagging strategies 65

Random and targeted tagging 65

Entrapment with transposons 67

Chromosomal engineering 69

5 Isolation of tagged genes 69

Proof that a gene has been tagged 69

Cloning a tagged gene 71

6 Reverse genetics strategies to identify mutants 75

Target selected gene inactivation 75

Sequencing of transposon flanking DNA to identify tagged genes 77

5 Cloning plant genes by genomic subtraction / Tai-ping Sun 83

2 The genomic subtraction technique 85

3 Deletion mutagenesis 85

4 Genomic subtraction protocol for Arabidopsis 87

Preparation of genomic DNA from wild-type and putative deletion mutants 87

Subtractive hybridization 90

Amplification and cloning of the DNA fragments after subtraction 91

Screening the clones corresponding to deletion mutations 93

Leak through background clones 95

Pilot experiments 96

5 Comparison of genomic subtraction to representational difference analysis (RDA) 97

Section 2 Gene Organization

6 Approaches to gene mapping / S. Grandillo, T. M. Fulton 101

2 Molecular mapping: an overview 102

Principles of linkage analysis 102

Molecular markers and genetic maps 103

Choice of mapping populations 104

Population size 105

Tagging genes with tightly linked markers 106

Gene mapping in two model plant species 107

Relationship between genetic and physical maps 110

3 Isolation of genomic DNA from plant tissue 111

4 Restriction fragment length polymorphism (RFLP) analysis 114

Restriction enzyme digestion and gel electrophoresis 115

Southern blotting 117

Preparation of nucleic acid probes 119

Hybridization analysis of Southern blots 122

5 Random amplified polymorphic DNA (RAPD) analysis 123

6 Principles of amplified fragment length polymorphism (AFLP) analysis 125

7 PFGE in physical mapping of plant genomes 127

Preparation of large fragment DNA from plant nuclei 128

Restriction digestion of agarose-embedded DNA 130

PFGE analysis 132

7 Arabidopsis YAC, BAC and cosmid libraries / Paul Muskett, Ian Bancroft, Caroline Dean 137

2 YAC libraries 137

Screening of YAC libraries by colony hybridization 139

Preparation of intact yeast chromosomes 141

3 BAC libraries 143

Construction of BAC libraries 144

Screening of BAC libraries using colony hybridizations 148

4 Cosmid libraries 152

Construction of a cosmid library using pCLD04541 153

8 In situ hybridization to plant chromosomes and DNA fibres / J. S. Heslop-Harrison, Trude Schwarzacher 159

2 Preparation of targets 160

Sources of material 160

Chromosome spread preparation 162

Extended DNA fibres 163

3 Probes and labels 165

The nature of probes 165

Cloned DNA sequences 165

PCR products 166

Total genomic DNA for chromosome and genome painting 166

Synthetic oligonucleotide sequences 167

Probe labelling and purification 167

Testing label incorporation and quality control of probes 168

Fluorochromes 168

4 The hybridization mixture, stringency and denaturation 169

Pretreatments of chromosome preparations 169

The hybridization mixture 170

Setting up the hybridization 172

Stringent washes 173

5 Fluorescent detection and visualization of hybridization sites 174

Detection of hybridization sites of indirect labels 174

Counterstaining and mounting 175

Visualization and imaging 176

Section 3 Library Screening and cDNA Isolation

9 PCR cloning approaches / Michiel Vandenbussche, Tom Gerats 179

2 RNA fingerprinting methods 180

Differential display 180

cDNA-AFLP 180

Obtaining full length cDNA clones: RACE-PCR 183

3 Multigene family cloning by the FamilyWalker method 183

The FamilyWalker strategy 184

Design of degenerate primers 187

Primer design 188

PCR optimization 190

Amplifying new Petunia hybrida MADS-box genes by the FamilyWalker method 193

10 Western and south-western library screens / Rhonda C. Foley, Karam B. Singh 205

2 Working with cDNA expression libraries 206

3 Preparation of nitrocellulose filters 208

4 Western cDNA library screening 209

Polyclonal and monoclonal antibodies 209

Detection systems used in western library screening 210

Characterization of clones isolated by western library screening 213

5 South-western screening approach 213

Preparing the DNA probe for south-western screening 214

Preparation of nitrocellulose filters for south-western screening 216

Using proteins that have been denatured/renatured for south-western screening 216

Screening the cDNA library for DNA binding proteins 217

Selection of positive phage 218

6 Characterization of clones isolated by south-western screening 219

11 Complementation cloning / E. Ann Oakenfull, James A. H. Murray 223

2 General requirements for complementation cloning 226

Choice of mutant strain and selection strategy 226

Choice of cDNA library and vector 230

Transformation efficiency 233

3 Preparation of cDNA libraries and mutant strains for complementation cloning 233

Amplification of plasmid cDNA libraries 233

Optimizing growth conditions of the mutant strain 236

4 Transformation and screening in E. coli 236

Transformation 236

Screening 236

5 Transformation and screening in S. cerevisiae 237

Transformation 237

Screening 239

6 Potential Problems 242.

Notes:

Rev. ed. of: Plant molecular biology. Oxford : IRL Press, 1988.

Includes bibliographical references and indexes.

ISBN:

0199638772

0199638764

0199638756

0199638195

OCLC:

49834342