Rapid purification of human DNA from whole blood for potential application in clinical 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 min, 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/chloroform 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 micrograms/mL, respectively. Within- and between-day CVs for recovery of DNA from pooled blood were 8% and 11% respectively. 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 apolipoprotein B gene.
Adeli
K
K
Department of Chemistry and Biochemistry, University of Windsor, Ontario, Canada.
Ogbonna
G
G
D001055:Apolipoproteins B / Q000235:genetics; D015139:Blotting, Southern; D002624:Chemistry, Clinical / Q000379:methods*; D004247:DNA / Q000097:blood / Q000302:isolation & purification*; D004252:DNA Mutational Analysis; D004262:DNA Restriction Enzymes; D004587:Electrophoresis, Agar Gel; D006801:Humans; D006868:Hydrolysis; D009203:Myocardial Infarction / Q000097:blood; D012150:Polymorphism, Restriction Fragment Length
D001055:Apolipoproteins B; D004247:DNA; D004262:DNA Restriction Enzymes