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DNA extraction from buffy coat
I'm looking to isolate DNA and eventually RNA from buffy coat. I receive whole blood in purple top vacutainers treated with EDTA. These tubes are centrifuged at 3000 rpm for 10 minutes to separate the whole blood into plasma buffy coat and rbc's. The plasma is drawn off with a transfer pipette to 2.0 ml microtubes and spun at 3000 rpm for 10 minutes to remove particulates, then transfered to cryovials and stored at -80.
I draw off the buffy coat and place it into a 15 ml conical tube filled with cold 10 mM tris 1mM EDTA. I let the tubes sit on ice for about 10 minutes then spin them at 3000 rpm for 10 minutes, pour off the supernatant and repeat with a few more 10 mL washes. A few rbc's persist but the majority are removed. The buffy coat is placed in a cryovial and stored at -80.
I'm using the fujifilm quickgene with various kits to isolate RNA and DNA from the frozen buffy coat.
I'm looking for suggestions on a more efficient method of obtaining buffy coat as well as extracting DNA.
|buffy , buffy coat , centrifuge , coat , dna , extraction , extraction or isolation|
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