Miniprep ! Can it bug you? - (Jul/01/2005 )
Its strange but I am facing problem with my miniprep. (Maniatis-alkaline lysis) After my ligation I transformed the cells(XL1Blue) which worked fine earlier. But now after miniprep I am not getting proper DNA band on gel. I see RNA, genomic dna and a light band in middle which could be plasmid dna. After pptn & centrifugation I get lot of sticky ppt on the walls of tubes which is not ready to dissolve in water with 40ug?ml RNase. What could be the problem?
3. Comp Cells??
Please help in finding the reason!!
This happened to me once. Check all reagents by taking a known plasmid like pBluescript and transform DH5alpha and do a mini prep on that. If it works, you know your reagents are okay.
Then redo the whole ligation/transformation thing using fresh made plasmid.
I know it sounds sorta simple.. but thats what I did to make it happen.