Green Fluorescent Proteins, Band 58Gulf Professional Publishing, 1999 - 386 Seiten This volume is an authoritative and comprehensive treatment of the approaches and techniques used for Green Fluorescent Proteins (GFP). The primary focus of this work is on research using biological systems. The volume covers all aspects of GFP, from its expression in different organisms to specific microscopic and data analysis methods. Key Features * Only volume on Green Fluorescent Protein research * Covers all aspects of GFP * Provides specific microscopic and data analysis methods * Discusses the design and construction of GFP fusion proteins * Covers GFP expression in animals, insects, plants, and microbes * Details procedures for time lapse imaging of living cells * Explains how to implement single molecule fluorescence detection with GFP * Discusses dual label GFP strategies for multicolor fluorescence * Presents fluorescence resonance energy transfer methods with GFPs * Details quantitative fluorescence imaging techniques * Extensively illustrated with color photographs |
Inhalt
Understanding StructureFunction Relationships in the Aequorea victoria | 19 |
Quantitative Imaging of the Green Fluorescent Protein GFP | 31 |
SingleMolecule Fluorescence Detection of Green Fluorescence Protein | 49 |
Targeting GFP to Organelles | 75 |
Cytoskeletal Dynamics in Yeast | 87 |
Analysis of Nuclear Transport in Vivo | 107 |
GFP Fusion Proteins as Probes for Cytology in Fission Yeast | 123 |
Applications of Fusion Proteins | 131 |
Postprocessing of Transfected Cells | 257 |
Overview | 258 |
Monitoring the Dynamics and Mobility of Membrane Proteins Tagged with Green Fluorescent Protein J LippincottSchwartz J F Presley K J M Zaal K... | 261 |
Brightness and Assessing Chimera Function | 262 |
Practical Guidelines for the Preparation and Imaging of GFPExpressing Cells | 264 |
Critical Parameters | 265 |
Analysis of TimeLapse Imaging Data | 268 |
Relating GFP Chimera Fluorescence to Actual Numbers of GFP Molecules | 270 |
GFP Fusions to a Microtubule Motor Protein to Visualize Meiotic | 153 |
GFP as a Cell and Developmental Marker in the Drosophila | 165 |
Visualizing GFP Expression | 175 |
Using TimeLapse Confocal Microscopy for Analysis of Centromere | 183 |
Visualization of LargeScale Chromatin Structure and Dynamics Using the lac Operatorlac Repressor Reporter System Andrew S Belmont Gang Li Gai... | 203 |
Overview of Methodology | 204 |
Construction of the lac Operator Repeat | 206 |
Manipulation of the lac Operator Repeats | 208 |
RepressorNLS and GFPRepressorNLS Constructs | 210 |
Gene Amplification and Cell Cloning | 212 |
Repressor Staining and Immunodetection of the lac Operator Repeat | 214 |
In Vivo Observation of GFPRepressor Localization | 215 |
Phototoxicity Issues | 216 |
Present Results and Future Directions | 218 |
References | 220 |
Centrosome Dynamics in Living Cells Aaron Young Richard Tuft Walter Carrington and Stephen J Doxsey | 223 |
Introduction | 224 |
Cloning and Expression of GFPPericentrin | 225 |
HighSpeed Microscopy | 228 |
Image Restoration by an Improved Deconvolution Method | 231 |
Imaging Centrosomes | 234 |
Postimaging Confirmation of Centrosome Integrity and Function | 235 |
References | 238 |
Transfections of Primary Muscle Cell Cultures with Plasmids Coding for GFP Linked to FullLength and Truncated Muscle Proteins Guissou A Dabiri ... | 239 |
Introduction | 240 |
Construction of GFPLinked Muscle Proteins | 241 |
Preparation of Embryonic Avian Cardiomyocytes and Skeletal Muscle Myoblasts | 243 |
Transfection of CrossStriated Muscle Cells with FullLength cDNA for Sarcomeric Proteins | 247 |
Microscopic Observations of Live Cells | 255 |
Fluorescence Recovery after Photobleaching FRAP | 271 |
Qualitative FRAP Experiments | 272 |
Quantitative FRAP | 275 |
Calculating D | 276 |
Fluorescence Loss in Photobleaching FLIP Using a Confocal Microscope | 278 |
Other Applications of Photobleaching | 280 |
Synchronous RealTime Reporting of Multiple Cellular Events Jeffrey D Plautz and Steve A Kay I Introduction | 283 |
Green Fluorescent Protein | 284 |
Luciferase | 285 |
Instrumentation and Techniques | 286 |
Future Directions | 290 |
References | 291 |
Visualizing Protein Interactions in Living Cells Using Digitized GFP Imaging and FRET Microscopy | 293 |
Ammasi Periasamy and Richard N Day I Introduction | 294 |
The Theory of FRET | 295 |
Review of the FRET Literature | 297 |
Why Use FRET Microscopy? | 298 |
Some Considerations for the Use of GFPs in FRET Imaging | 301 |
Some Considerations for Designing a FRET Imaging System | 303 |
The Practical Application of FRET to Visualize Protein Protein Interactions | 307 |
Overview and Conclusion | 309 |
References | 310 |
Flow Cytometric Analysis and FACS Sorting of Cells Based | 315 |
Imaging Gene Expression and Protein Dynamics | 343 |
Imaging Systems | 349 |
42 | 366 |
| 369 | |
| 375 | |
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