I've been struggling to get consistent results from my Pichia transformations. I've tried both heat shock and electroporation protocols. I suspect that part of my problem is that the cultures tend to flocculate, which makes it very difficult to accurately measure OD600, so its hard to know when to harvest cultures for making competent cells.
I'm working with the Pichia Pink strains from Invitrogen, and growing my cultures for making competent cells in standard YPD broth. I've been trying to measure OD600 in a plate reader.
Can anyone suggest a solution to this problem? I'm sure I'm not the only one to encounter it.
Hi there,
Light sonication of the sample right before measurement should be OK.
I've never had Pichia strains flocculating in standard YPD. I'm curious, is this a common occurrence with the Pichia Pink system?
Thank you for the advice. I'll try EDTA.
Alejandro Martin - In my experience the Pichia pink strains often (but not always) flocculate in YPD, BMGY and BMMY. Whether or not any particular culture will flocculate seems almost impossible to predict. If it happens it generally seems to be in late log or stationary phase, rather than earlier on.
I should add that I'm not very experienced at working with yeast. Apart from brewing the occasional beer at home I've only worked with the Pichia pink strains, so I have no basis for comparison.
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