Genotyping of DNA samples isolated from formalin-fixed paraffin-embedded tissues using preamplification.
DNA isolated from formalin-fixed paraffin-embedded (FFPE) tissue is often fragmented and cross-linked and is therefore difficult to genotype. To enable this source of DNA for genotyping analysis using Taqman probes, we tested whether enrichment of the target genes would increase the amount of available DNA. For enrichment of the target genes, we used preamplification by means of diluted Taqman assays. To establish the appropriateness of preamplification, we used DNA extracted from paraffin-embedded tissue and compared the genotyping results of a series of single nucleotide polymorphisms assessed in DNA samples with and without preamplification. In a subset of patients, DNA was isolated from both blood and FFPE tissue to test the reliability of genotyping results derived after preamplification. We found an increase in call rate after preamplification and a convincing concordance in genotype. Based on our findings, we can safely conclude that preamplification of DNA isolated from paraffin-embedded tissue is a valuable and reliable method to optimize genotyping results.
Baak-Pablo
Renee
R
Department of Clinical Pharmacy and Toxicology, Leiden University Medical Center, P.O. Box 9600, 2300 RA Leiden, The Netherlands.
Dezentje
Vincent
V
Guchelaar
Henk-Jan
HJ
van der Straaten
Tahar
T
D004247:DNA / Q000032:analysis* / Q000302:isolation & purification; D056915:DNA Copy Number Variations; D005557:Formaldehyde; D005838:Genotype; D006801:Humans; D021141:Nucleic Acid Amplification Techniques; D016612:Paraffin Embedding
D005557:Formaldehyde; D004247:DNA