- 作 者：（美）Dan Burke等著
- 出 版 社：北京：清华大学出版社
- 出版年份：2002
- ISBN：7302050961
- 标注页数：205 页
- PDF页数：221 页
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Experiments 1
Ⅰ.Looking at Yeast Cells 1
Ⅱ.Isolation and Characterization of Auxotrophic,Temperaturesensitive,and UV-sensitive Mutants 7
Ⅲ.Meiotic Mapping 17
Ⅳ.Mitotic Recombination and Random Spore Analysis 29
Ⅴ.Transformation of Yeast 39
Ⅵ.Synthetic Lethal Mutants 49
Ⅶ.Gene Replacement 55
Ⅷ.Isolation of ras2 Suppressors 73
Ⅸ.Manipulating Cell Types 79
Ⅹ.Isolating Mutants by Insertional Mutagenesis 89
Ⅺ.Two-hybrid Protein Interaction Method 97
Techniques and Protocols 103
1.High-efficiency Transformation of Yeast 103
2.“Lazy Bones”Plasmid Transformation of Yeast Colonies 107
3.Yeast DNA Isolations 109
A.Yeast DNA Miniprep(40 ml) 109
B.Yeast DNA Miniprep(5 ml) 110
C.A Ten-minute DNA Preparation from Yeast 111
D.Yeast Genomic DNA:Glass Bead Preparation 113
4.Yeast Protein Extracts 115
5.Yeast RNA Isolation 117
6.Hydroxylamine Mutagenesis of Plasmid DNA 121
7.Assay of β-Galactosidase in Yeast 123
8.Plate Assay for Carboxypeptidase Y 127
9.Random Spore Analysis 129
10.Yeast Vital Stains 131
11.Yeast Immunofluorescence 133
12.Actin Staining in Fixed Cells 137
13.PCR Protocol for PCR-mediated Gene Disruption 139
14.Yeast Colony PCR Protocol 141
15.Measuring Yeast Cell Density by Spectrophotometry 143
16.Cell Synchrony 145
17.Chromatin Immunoprecipitation 149
18.Flow Cytometry of Yeast DNA 155
19.Logarithmic Growth 157
20.EMS Mutagenesis 159
21.Tetrad Dissection 161
22.Making a Tetrad Dissection Needle 165
23.Picking Zygotes 167
24.Determining Plating Efficiency 169
Appendices 171
A.Media 171
B.Stock Preservation 183
C.Yeast Genetic and Physical Maps 185
D.Templates for Making Streak Plates 193
E.Electrophoretic Karyotypes of Strains for Southern Blot Mapping 195
F.Strains 197
G.Counting Yeast Cells with a Standard Hemocytometer Chamber 199
H.Tetrad Scoring Sheet 201
I.Trademarks 203
J.Suppliers 205