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Submitted by [[User:Smoore|Sean Moore]] | Submitted by [[User:Smoore|Sean Moore]] | ||
==Getting The Most Out Of Your Bugs== | ==Getting The Most Out Of Your Bugs== | ||
Native lysis is a staple protocol in practically every biochemistry lab, yet there is significant variability in the procols I have read. The intention is to liberate the guts of E. coli without disturbing the native conformations of the biomolecules. Using a French pressure cell you can | Native lysis is a staple protocol in practically every biochemistry lab, yet there is significant variability in the procols I have read. The intention is to liberate the guts of E. coli without disturbing the native conformations of the biomolecules. Using a French pressure cell you can sheer the cells open, this is a great way to lyse cells. Unfortunately, it is impractical for multiple samples or small-volume samples that don't fit in the pressure cell. Besides, the machine is scary, lysis can be variable, and cleanup is a hassle. Chickens and bacteriophage have evolved great a way of opening E. coli using enzymes. Most commercial lysozymes are free from proteolytic activity and can be added in large amounts. Be careful, if your protein is ~14 kDa you may inadvertantly purify the added lysozyme instead of your target protein. Don't laugh, it's happened more than once and I have met people who solved the crystal stucture of lysozyme by mistake. | ||
===Hen Egg White Lysozyme=== | ===Hen Egg White Lysozyme=== |
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