Recovery of genomic DNA from residual frozen archival blood clots suitable for amplification and use in genotyping assays.
A greatly neglected source of DNA potentially useful for genetic or forensic studies is the clot remaining from blood samples collected for serum chemistry measurements. We have investigated the utility of residual clots remaining from venipunctures collected for California's Expanded Maternal Serum Alpha-Fetoprotein Screening Program. We report a protocol based on the salting out method for the extraction of DNA from samples which have been archived and frozen for up to 2.5 years. As much as 57 microg of high-quality DNA can be obtained from a 2-ml clot as determined by PicoGreen (Molecular Probes, Inc., Eugene, OR) fluorescence measurements. Quality of the purified DNA was evaluated by its ability to serve as template in polymerase chain reaction (PCR) amplifications, using primers that flank the polymorphic regions of six genes of pharmacogenetic interest distributed throughout the human genome. Sizes of the gene regions successfully amplified range from 215 bp to 2064 bp, using as little as 10 ng of template DNA. Because many genotyping protocols routinely recommend the design of amplicons in the 100-200 bp range, and 10-50 ng of template, we conclude that the clot remaining after serum has been removed from blood collected for serum chemistry measurements can serve as a reliable source of DNA for genotyping studies.
Iovannisci
David M
DM
Children's Hospital and Research Center Oakland, Oakland, California 94609, USA. Diovannisci@chori.org
Ha
Thanh T
TT
Shaw
Gary M
GM
D018070:Biological Specimen Banks; D001769:Blood; D015925:Cryopreservation / Q000379:methods; D004247:DNA / Q000737:chemistry / Q000302:isolation & purification*; D016172:DNA Fingerprinting / Q000379:methods; D015894:Genome, Human; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods
D004247:DNA