Evaluation of GenoType MTBDRplus by Use of Extracted DNA from Formalin-Fixed Paraffin-Embedded Specimens.
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ABSTRACT:
SUBMITTER: Schaumburg F
PROVIDER: S-EPMC5654916 | biostudies-literature | 2017 Nov
REPOSITORIES: biostudies-literature
Publications
Evaluation of GenoType MTBDR<i>plus</i> by Use of Extracted DNA from Formalin-Fixed Paraffin-Embedded Specimens.
Schaumburg Frieder F Peters Georg G Wardelmann Eva E Becker Karsten K Sperveslage Jan J
Journal of clinical microbiology 20170906 11
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Project description:Single-nucleotide polymorphisms (SNPs) can be assayed using DNA isolated from archival formalin-fixed, paraffin-embedded (FFPE) samples, making retrospective pharmacogenetic studies possible. In this study, we describe methods that significantly increase the number of SNP determinations possible using FFPE samples. Quantifying the amount of DNA amenable to PCR (amplification-quality DNA, AQ-DNA) allows a significant reduction in the amount of sample required for Taqman-based SNP assays. Optimizing AQ-DNA input increases PCR amplification efficiency and SNP determination accuracy. DNA was extracted from 39 FFPE tumor sections and matched tumor and stromal cores, which were of the type used to generate tissue microarrays. Sections and tumor cores yielded sufficient AQ-DNA for more than 1000 SNP determinations. Seven SNPs were assessed following individual assay optimization for minimal AQ-DNA. Genotypes from tumor cores for single SNPs were 92.3-100% concordant with those obtained from sections. Using these methods, the number of SNP genotypes that can be determined from single FFPE samples is greatly increased expanding the genetic association studies possible from limited archival specimens. The use of tumor cores is of particular importance as the harvesting of tumor cores has minimal impact on the utility of the donor blocks for other purposes.
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