Optimization of extraction and PCR of DNA from frozen serum

A variety of biological materials have boon used as a DNA source for PCR. DNA extracted from bnffy coats is mot commonly used for genetic studies. Howevoi, it is not always possible to obtain a huffy coat from a subject for DNA extraction. In addition, many laboratories have large banks of frozen sera stored on specific patient populations that could be used for genetic studies. These observations prompted us tooptimi/o DNA extraction and amplification from fro/en serum. Initially, thirteen DNA extraction methods were evaluated in a srn<ill population. Four of these methods were chosen for further study based on A260:280, DNA recovery and ease of method. Froxen sera from 110 pro.statr- cancer patients were extracted by boiling, Chelex 100 incubation followed by boiling, a QIAamp blood kit or an Ka.y-DNA kit. DNA recovery was calculated by spectrophotometry arid fluorescence assays. Depending on the extraction and quantitation method, the DNA recovered was 12-57% of i he DNA present in native serum. DNA extracted by QIAamp or Easy-ON A had an A260:280 of 1.2-1.9; the other method.-, had a median ratio of 0.8. A triniitleotide repeat region in the androhen receptor was amplified to determine t lie suitability of the DNA in PCH. lioilinp, methods were unreliable with only 10 20CX of the DNAs amplifiable; using QIAamp or Kasy-DNA. 77% were amplified. Fasy DNA uses organic solvents and is time consuming; QIAamp is based on a silica column and requires thiiiv minutes. From our results, the optimal choice was the QIAamp procedure. II there was no amplification by QIAamp. then Fasy-DNA was performed. Using, this approach. 84.3% of the DNAs extratted from fro/en serum wre amplified.