Genotyping rs61753984 (R34X) and rs62643623 (Q218X) in Von Willebrand disease patients by tetra ARMS-PCR

Von Willebrand disease (VWD) is the most common inherited bleeding disorder around the world. Large size of the gene and heterogeneous nature of its mutations, make it difficult to determine the exact nature of mutations in VWD. Some missense mutations are common in VWD and can be assessed as first step of determining the genetic cause of the disease.

Forty four unrelated patients categorized as VWD type 3 were included in study. Tetra ARMS-PCR was applied to determine the presence of the point mutations rs61753984 and rs62643623. Then ARMS-PCR was performed with the same primers. In cases with mutant patterns, Sanger sequencing was used for conformation of results.

Two patients showed two bands of 248bp and 181bp in rs61753984 (R34X), which corresponds to homozygous mutant genotype. In other patients, homozygous pattern with 248bp and121bp bands was observed. In rs62643623 (Q218X) position, one patient with two bands of 378bp and 260bp showed homozygous mutant genotype while other patients showed homozygous wild genotype (378bp and 168bp bands). The results of ARMS-PCR was the same as tetra ARMS-PCR. Finally, the results of tetra ARMS-PCR were confirmed by Sanger sequencing.

In present study we used tetra ARMS-PCR for detection of point mutations, while in most studies PCR-PFLP has been used. We found that tetra ARMS-PCR can be used as a simple, cost effective and reliable method. If these mutations are not found, advanced molecular methods such as sequencing should be applied.