Czech

2

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NOW ( C7230 Fluorescence methods

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The course is taught: in blocks.

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Spectral characterization of proteins and nucleic acids
Accurate determination of concentration of nucleic acids and proteins by fluorescence and absorption spectroscopy (colorimetry)
Effects of pH and temperature on spectral properties of fluorophores
Measurement of intrinsic fluorescence of BSA, tryptophan, and thyroxin
Preparation of fluorescently labeled DNA
Spectroscopic characterization of fluorescently labeled DNA
Visualization of macromolecules after electrophoretic separation
Use of resonance energy transfer (FRET) for detection of DNA hybridization
Fluorescence quenching after relaxation of loop structure of fluorescently labeled DNA
Real-time PCR – Detection of DNA amplification
Fluorescence microscopy (fluorescence in situ hybridization)
Fluorescence anisotropy – Study of protein binding to a fluorescently labeled DNA

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The main objective of the practical training course is to acquire practical experience with modern spectroscopic methods employed in the analysis of protein-protein and protein-DNA interactions. From the methodical approaches, the main emphasis is put on highly sensitive fluorescent methods and their practical use in biological laboratories. The practical one week course, complementing lectures of Bi7230, enables trainees to corroborate their theoretical knowledge about methodical principles using modern instrumentation of research laboratories. Training is focused on the effective development of skills required for application of specialized techniques. These techniques have become indispensable for recent tasks of basic and applied research in today’s interdisciplinary concept of biological sciences.

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Lakowicz J.R.: Principles of Fluorescence Spectroscopy. Third Edition, Springer + Business Media, New York, 2006 (CS)

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