DNA Southern blot with electric current

Abir Mukherjee abrim at casbah.acns.nwu.edu
Thu Nov 26 21:37:27 EST 1998


In article <73cpbc$eq3$1 at nnrp1.dejanews.com>, gigles at fvet.uba.ar wrote:

> I wish to know if thats possible, how can I do a Southern using electric
> current I need to know about the buffer, volts and amperes that I must use.
> Thanks.
> 

I have done this only a couple of times. I had run a 5% acrylamide gel in
1XTBE. I transferred the DNA onto nylon membrane using our western
transfer apparatus, in TBE buffer and at 50 volts for 1.5 hours. After
transfer I UV-crosslinked the membrane and then denatured the DNA on the
membrane by allowing the membrane to incubate on denaturation buffer
soaked whatman paper for 5 min. This was followed by incubation on whatman
papers soaked with neutralization buffer and 2XSSC. I was looking for a
fragment of about 250 bp. It actually worked!

Good luck



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