Dataset Information

Large‐scale screening for factor V Leiden (G1691A), prothrombin (G20210A), and MTHFR (C677T) mutations in Greek population

ABSTRACT: Abstract

Background and aims

To provide a fair estimate of the prevalence of factor V Leiden (FVL) (G1691A), prothrombin (G20210A), and MTHFR (C677T) mutations in the Greek population.

Methods

We genotyped a representative sample of 974 apparently healthy Greek adults by the method of real‐time PCR and we calculated the allele frequencies of factor V Leiden (FVL) (G1691A), prothrombin (G20210A), and MTHFR (C677T) mutations. In addition, we determined the frequency of co‐occurrence of FVL (1691A) and prothrombin (20210A), FVL (1691A) and MTHFR (677T), prothrombin (20210A) and MTHFR (677T) mutations.

Results

Τhe career frequencies of FVL (1691A), prothrombin (20210A), and MTHFR (677T) alleles were 7.5%, 4.5%, and 49.3% while the allele frequencies were 4%, 2.25%, and 39.5%, respectively. The coexistence of the allele frequencies combinations of two, FVL (1691A) and Prothrombin (20210A), FVL (1691A) and MTHFR (677T), prothrombin (20210A) and MTHFR (677T) was found in 1 (0.9%), 29 (3.5%), and 22 (3%) samples, respectively. Triple heterozygous carriers were not found.

Conclusion

Allele frequencies of the two (FVL and MTHFR) mutations are higher compared with published data. The large sample size of our study enhances the validity of our results and suggests a biological affinity of Greek population with Southern Italian populations.

SUBMITTER: Raptopoulou A

PROVIDER: S-EPMC9284178 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Factor V Leiden and prothrombin G20210A are clinically relevant genetic risk factors for venous thrombosis. Analysis for both mutations is increasingly being performed on patients exhibiting hypercoagulability. The goal of the current study was to evaluate the performance of primer-engineered multiplex polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) for the simultaneous detection of factor V Leiden and prothrombin G20210A. Primer-engineered multiplex PCR-RFLP methods for the detection of factor V Leiden and prothrombin G20210A from the medical literature were reviewed. A modified method was optimized in which both mutations generate HindIII RFLPs and the prothrombin amplicon contains an invariant HindIII recognition site to assess the completeness of endonuclease digestion. Digested amplification products were analyzed by agarose gel electrophoresis in a single gel lane and visualized by ethidium bromide. Primer-engineered multiplex PCR-RFLP was used to analyze 205 human genomic DNA samples whose factor V Leiden genotypes had been previously determined by MnlI PCR-RFLP. Complete concordance for factor V Leiden genotypes was observed between the two methods in the 205-sample cohort comprising 139 wild-type, 62 heterozygous mutant, and four homozygous mutant individuals. For prothrombin G20210A, primer-engineered multiplex PCR-RFLP identified 196 wild-type and nine heterozygous mutant individuals in the 205-sample cohort. To independently verify prothrombin genotypes, the nine heterozygous mutants and an additional 11 wild-type patient samples (representing 10% of patient samples) were subjected to DNA sequencing. Complete concordance was observed between DNA sequencing and primer-engineered multiplex PCR-RFLP results. In further validation, 123 of the DNA samples consisting of four heterozygous mutant and 119 wild type individuals were genotyped with the Invader Assay for Factor II (prothrombin G20210A). Results showed 100% concordance between the Invader Assay and primer-engineered multiplex PCR-RFLP. A primer-engineered multiplex PCR-RFLP based on single restriction endonuclease digestion has been evaluated and shown to simultaneously and accurately detect factor V Leiden and prothrombin G20210A mutations. The method is robust and readily adaptable to the clinical molecular diagnostic laboratory.