I recently used the method of Witman (JCB, 1978, 76, 729) to isolate and
reactivate axonemes from C. reinhardtii and found the yield of axonemes
purified by this method to be very low.
I am wondering if anyone with experience purifying/reactivating axonemes
might lend some advice. I started with 400 mL of culture grown
synchronously using a light/dark cycle. Cells were clearly swimming
before starting the procedure. I was surprised not to see any axoneme
pellet during the high g spins, but attributed this to isolating only a
small amount of protein. Perhaps these spins were too slow/short?
Any advice/troubleshooting would be most welcome.
Thanks in advance,
Doug
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