Sauer:Lysing E. coli with Lysozymes: Difference between revisions
No edit summary |
|||
| Line 4: | Line 4: | ||
===Hen Egg White Lysozyme=== | ===Hen Egg White Lysozyme=== | ||
I prepare the lysozyme fresh each time from lyophilized powder. Pre-warm the bottle to prevent moisture from condensing in the bottle when it's opened. Pure B-Per doesn't seem to allow it into solution and it remains mostly as an inactive precipitate. So I make a ~10 mg/mL stock in some buffer off my shelf (I usually use 50 mM bis-Tris, pH 6.5). Then I dilute from this to 0.1 to 0.01 mg/mL in the B-Per lysis solution. To save time, I use a pipette tip and lift out dried lysozyme horizontally from the bottle, I then place a microfuge tube over the tip and shake off the powder. I measured the before and after weight of several tubes and found that I can make pretty close to a 10 mg/mL solution by just adding 400 uL of buffer to the tube. So, I no longer weigh out lysozyme each time, I just assume that my tip carries about 4 mg of protein. | |||
===Intracellular Lysozymes=== | ===Intracellular Lysozymes=== | ||
Revision as of 14:33, 24 January 2007
Submitted by Sean Moore
Getting The Most Out Of Your Bugs
Hen Egg White Lysozyme
I prepare the lysozyme fresh each time from lyophilized powder. Pre-warm the bottle to prevent moisture from condensing in the bottle when it's opened. Pure B-Per doesn't seem to allow it into solution and it remains mostly as an inactive precipitate. So I make a ~10 mg/mL stock in some buffer off my shelf (I usually use 50 mM bis-Tris, pH 6.5). Then I dilute from this to 0.1 to 0.01 mg/mL in the B-Per lysis solution. To save time, I use a pipette tip and lift out dried lysozyme horizontally from the bottle, I then place a microfuge tube over the tip and shake off the powder. I measured the before and after weight of several tubes and found that I can make pretty close to a 10 mg/mL solution by just adding 400 uL of buffer to the tube. So, I no longer weigh out lysozyme each time, I just assume that my tip carries about 4 mg of protein.
