Need advice on soybean protoplast

Nanfel Xu nxu at
Wed Nov 2 20:35:43 EST 1994

      I am trying to isolate protoplast from cotyledons of
germinated soybean seeds.  Some of the protoplast can be liberated
by 0.1% pectolyase and 3% cellulase (both from Sigma).  But I have
two problems.  First, the liberated protoplasts become brown (and
presumably died) after an hour or so, while waiting for more
protoplasts to be released.  Also, the work of cellulase seems to
be quit slow.  By 12 h, almost all the cotyledon pieces are broken
into single cells, but the proportion of protoplast is small.  Has
anybody encountered this kind of problems before?  What can I do to
improve the yield and quality of these protos?  Thank you.

Nanfei Xu

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