Chemistry Laboratories

A simple, rapid method for isolating human DNA has been developed, which can be routinely used in clinical chemistry laboratories. The entire procedure takes less than 90 mm, and as many as 12 blood samples can be handled in one cycle. One milliliter of EDTA-treated blood is lysed and centrifuged to yield a nuclear fraction. The nuclear pellet is treated with sodium dodecyl sulfate/urea and phenol/chloro-form to remove contaminating proteins, then the crude DNA extract is purified by use of a Sephadex G-25 spin-column. Typical 260 nm/280 nm absorbance ratio (used to assess purity) and yield for DNA so purified were 1.84 and 24.5 zg/mL, respectively. Within- and between-day CVs for recov-ery of DNA from pooled blood were 8 % and 11%, respec-tively. Such DNA preparations were found quite suitable for digestion by a variety of restriction endonucleases and for restriction fragment length polymorphism analysis. We are using this method to isolate DNA from whole blood of myocardial infarction patients for studies on the apolipopro-tein B gene. Additional Keyphrases: gene probe. apolipoprotein B electrophoresis, agarose gel. recombinant DNA techniques Potential applications of recombinant DNA techniques in clinical chemistry laboratories are becoming increas-ingly evident, especially for diagnostic assays for many genetic and other diseases (1-3). Currently, DNA diagnos-tic assays are available for only a few rare disorders, but the field is rapidly expanding and new assays are being developed (4). Most DNA diagnostic tests involve extrac-tion of DNA from human leukocytes, traditionally by time-consuming and cumbersome procedures (5, 6). The most popular method for this involves digestion with pro-teinase K, takes about two days, and requires a large volume of blood (10-50 mL) (5). Such techniques are obviously not well-suited for use in routine clinical labora-tories. A faster, simpler method is required for the intro-duction of DNA analysis to the clinical chemistry labora-tories. We have developed a simple spin-column method for purification of DNA from human whole blood. Its simplicity and rapidity makes this method suitable for routine appli-cations in DNA analysis