Use of whole genome amplification to rescue DNA from plasma samples.
While DNA of good quality and sufficient amount can be obtained easily from whole blood, buccal swabs, surgical specimens, or cell lines, these DNA-rich sources are not always available. This is particularly the case in studies for which biological specimens were collected when genotyping assays were not widely available. In those studies, serum or plasma is often the only source of DNA. Newly developed whole genome amplification (WGA) methods, based on phi29 polymerase, may play a significant role in recovering DNA in such instances. We tested a total of 528 plasma samples kept in storage at -40 degrees C for approximately 10 years for 8 single nucleotide polymorphisms (SNPs) using the 5' exonuclease (TaqMan) assay. These specimens yielded undetectable levels of DNA following extraction with an affinity column but produced an average 52.7 microg (standard deviation of 31.2 microg) of DNA when column-extracted DNA was used as a template for WGA. This increased the genotyping success rate from 54% to 93%. There were only 3 disagreements out of 364 paired genotyping results for pre- and post-WGA DNAs, indicating an error rate of 0.82%. These results are encouraging for expanding the use of poor DNA resources in genotyping studies.
Lu
Yanhui
Y
International Agency for Research on Cancer, Lyon, France.
Gioia-Patricola
Lydie
L
Gomez
Jorge Vivas
JV
Plummer
Martyn
M
Franceschi
Silvia
S
Kato
Ikuko
I
Canzian
Federico
F
D000483:Alleles; D017103:Bacillus Phages / Q000378:metabolism; D001709:Biotechnology / Q000379:methods; D004247:DNA / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers / Q000494:pharmacology; D005821:Genetic Techniques; D015894:Genome, Human; D005838:Genotype; D006801:Humans; D021141:Nucleic Acid Amplification Techniques; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D015203:Reproducibility of Results; D013048:Specimen Handling; D013696:Temperature
D017931:DNA Primers; D004247:DNA