Aeration is critical for successful expression of recombinant proteins in yeast. I was wondering if there is any approach for fast small scale screening of expression rate.
Hello and Good day....
My research previously deal with a lot of sample....around 2000...my sample is recombinant e.coli...so i need plate that can growth all of my sample in shaking condition...i used one plate containing 20 well...each well can load around 1 or 2 ml of culture...it produce good growth for my bacteria..i think is suitable for your case also..
Hello Roy,
I cultured Pichia in a 96 deep-well plate for several high throughtput screenings. This type of plates allow you a better aeration than the typical 96-well plates. So, I could expect that your 24-well plate can run.
FEMS Yeast Res. 2004 Nov;5(2):179-89.
Reliable high-throughput screening with Pichia pastoris by limiting yeast cell death phenomena.
Weis R1, Luiten R, Skranc W, Schwab H, Wubbolts M, Glieder A.
Hi Roy,
You can indeed use multiwell plates for the Pichia expression studies. Aeration is certainly important, and if you have sub-optimal aeration your expression levels will be lower, but should still be detectable.
There are other options though - if all you want to do is to screen for expression and don't plan to purify the protein until you scale up you can use a membrane based screening method. I've previously done this by replica plating the colonies to a PVDF membrane (you can get discs of these that fit into plates) and growing them on MG plates for a couple of days, then transferring the membrane to an MM plate to induce expression. The cells are then rinsed off the membrane and the protein of interest is detected using appropriate antibodies. I've found that this method has allowed me to screen far more colonies that growth in liquid culture.
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