A set of five microsatellite markers linked to F8 gene can detect haemophilia a carriers across India

Abstract

Haemophilia A (HA) is an X-linked recessive bleeding disorder, primarily because of defects in the 186-kb long factor VIII gene (F8) affecting 1–2 men per 10 000 worldwide. Available markers for carrier detection are not effective in all populations, especially in India. In this study, we have chosen a set of five microsatellite markers, namely, DSX9897, DSX1073, intron 1 (GT)_n, intron 22 (CA)_n and intron 25 (CA)_n, in and around the F8 gene to achieve better sensitivity for carrier detection. Each marker locus has been PCR amplified in the individual DNA samples using fluorescent markers followed by genotyping experiment in automated sequencer. Genotype calls have been made by GeneMapper Software (version 4). Allele frequency of each microsatellite marker was calculated manually. Heterozygosity was determined by counting the heterozygotes in the female subset. We have shown that in 253 normal individuals from 20 different ethnic groups of India, the heterozygosity for the markers ranged from 0.25 to 0.54; and for the entire subset of 102 female samples we could successfully discriminate between the two X-chromosomes using these five markers. These markers could also discriminate between the two X-chromosomes for each of 39 obligate carriers included in this study. In conclusion, this panel of five markers around the F8 locus can be used for carrier detection of HA with higher sensitivity across India for families affected with the disease.