Journal article
Direct blood PCR: TaqMan-probe based detection of the venous thromboembolism associated mutations factor V Leiden and prothrombin c.20210G>A without DNA extraction
Clinica chimica acta
Jan 2019
Abstract
BACKGROUND
Practically, the initial step of genetic analysis is the extraction of DNA from blood or other cells, which is often time consuming and cost-intensive. We aimed at establishing a real-time PCR protocol for the detection of the venous thromboembolism associated mutations factor V Leiden (F5 c.1691G>A; p.R506Q) and prothrombin (F2) c.20210G>A from whole blood, without DNA extraction.
METHODS
F5 c.1691G>A (p.R506Q) and F2 c.20210G>A mutations were determined in 205 EDTA anti-coagulated whole blood samples from patients who underwent routine clinical genotyping using the DirectBlood Genotyping PCR Kit (myPOLS Biotec, Konstanz, Germany) together with in-house developed TaqMan primer-probe assays.
RESULTS
Validity score values of genotype calls using whole blood were similar and did not significantly differ compared to those using genomic DNA as substrate in PCR. Mutation analysis of 205 whole blood samples showed a negligible PCR dropout rate (one in 410 reactions) and were in 100% concordance with results obtained by conventional genotyping.
CONCLUSION
We successfully established a robust and valid real-time PCR protocol for the detection of the venous thromboembolism associated mutations F5 c.1691G>A (p.R506Q) and F2 c.20210G>A directly from whole blood.
Metrics
5 Record Views
7 citations in Web of Science
8 citations in Scopus
Details
- Title
- Direct blood PCR: TaqMan-probe based detection of the venous thromboembolism associated mutations factor V Leiden and prothrombin c.20210G>A without DNA extraction
- Creators
- Kathrin Geiger - Vorarlberg Institute for Vascular Investigation and TreatmentAndreas Leiherer - Private University in the Principality of LiechtensteinEva-Maria BrandtnerPeter Fraunberger - Vorarlberg Institute for Vascular Investigation and TreatmentHeinz Drexel - Drexel University, College of MedicineAxel Muendlein - Vorarlberg Institute for Vascular Investigation and Treatment
- Publication Details
- Clinica chimica acta
- Publisher
- Elsevier
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- College of Medicine
- Other Identifier
- 991021860708304721
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- Collaboration types
- Domestic collaboration
- International collaboration
- Web of Science research areas
- Medical Laboratory Technology