Difficulty Isoalting DNA From Adipose Tissue - (Apr/05/2007 )
I am new to the boards, but in searching have found quite a bit of valuable information.
Currently I am using the Qiagen DNeasy Tissue Kit to isolate adipose, skeletal muscle, and brain tissue. In particular I am having poor yields with the adipose tissue of less than 0.2ug/mg of tissue while being careful to not overload the column (less than 25mg of tissue). I have been digesting the tissue overnight at 57C and then adding RNAase at RT, and spinning to float to the lipids. I take the portion underneath the floating lipids, but I am not positive some lipid might be coming with and eventually ended up on the column. Do you think that this could lead to the small yield. My 260/280 ratio is about 1.6-1.7, also a little low, but I feel that if I can increase my yield the ratio will improve. Any thoughts/suggestions would be much appreciated.
You might try extracting the adipose tissue with chloroform to remove excess lipids as a first step.