530L_notes_lecture6

530L_notes_lecture6 - X. Cell lysis A. Methods: 1....

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X. Cell lysis A. Methods: 1. Sonication High frequency Sound 2. Blender (mechanical) 3. French Press High pressure, long tube, and narrow aperture 4. Lysozyme (enzymatic) Enzyme that breaks down or degrade bacterial cell wall 5. Freeze/Thaw Rapidly freezing and thawing cells ***Depending on what type of proteins you are trying to collect, the cell lysis method varies. In this lab we chose to use lysozyme and freeze/thaw mainly for economic reasons. It works quite well. B. How does freeze thaw work? 1. Expansion and contraction By rapidly changing the temperature, make cell swell and shrink – alternating between dry ice and warm water bath. lysozyme’s role and bacterial cell wall? IMPORTANT Cell wall gives bacterial their structure. Addition of lysozyme breaks down this cell wall. Then, treat cells meanly: Freezing and thawing makes their cell membrane to break down. This combination causes the cells to lyse and dump their cellular constituents into the re- suspension buffer. C. Comparison—extracting DNA vs Proteins from cells 1. Methodology: It is far less aggressive than extracting DNA because proteins may denature. Since proteins are held together by weak forces, less aggressive method is used
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530L_notes_lecture6 - X. Cell lysis A. Methods: 1....

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