I use PEG for gene transformation of petunia protoplasts.

I use PEG4000...

When washing is done, the cells aggregate and eventually die.

I guess it's because the concentration of the cells are too high

they get crushed while centrifuging..

But I saw pictures of cells exhibiting GFP with high density.

This time, it was about 233 x 10(4) cells just before I treated them with PEG.

How can I prevent them from aggregation?

I really need to keep the cells alive so I can see fair efficiencies of transformation..

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