Help : Rat embryo fibroblasts and transformation

[Jun Chang]

Hi,
I need some help from experts about primary cell culture.
I'm studying the transforming potentials of viral genes using
rat embryo fibroblasts (REF), but I have some problems.
First, I have transfected E1A- and Ha-ras-expressing plasmids
to one million REF cells isolated from 14-day-old embryo as a
control, and obtained about 80 foci per 100mm dish. But next time
I did the same control experiment with REF from differnt rat, and no
foci at all! Is there some variations between different rats of 
same species? I used Sprague-Dawley rats.
Second, should I use supercoil plasmids or linearized ones? Till now
I used supercoils. In many reference papers, there is no comment about
the state of plasmids.
If you know about these questions, please answer me.
I would appreciate very much.







 	This is written by Chang Jun (Jeje).		   
 	E-mail : Jeje at sws1.postech.ac.kr		   
 	Pohang Univ. Sci. & Tech.			   
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