I need to transform a yeast with a vector. in yeast, plasmids need to be linearized prior to transformation. I am trying to purify digested dna but i ran out of gel extration kit. Any ideas? can i use a plasmid miniprep kit spin column instead? (i am afraid that linear dna to be alkaline degraded)
Anyway: Can i transform with the digestion (heat inactivated) without purifying it?
Thanks
I´m recomender to use the kit DNA clean and concentrador tm-5 of Zimo Research, the protocol is very fast, in 10 minutes you have you DNA clean and ready to transformation protocol. For other hand you can use phenol for clean your DNA without using a comercial kit. In my case the kit works very well.
Hi
Phenol:chloroform based purification is an established method in absence of a kit.
I have used the digestion to transfect mamalian cells, I think if you inactivate the restriction enzyme it will be OK.
OK! thank you guys!! I will try something of above before my kit arrives...
Guillem,
I have always used linearized DNA directly from a restriction reaction to transform yeast. Attempting to purify the linearized DNA would lead to the loss of a lot of DNA, unnecessarily, and none of the other components will hurt the transformation.
Patricia - thanks for pointing out that purification isn't necessary for transfecting mammalian cells either!
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