A rapid multifluorescent polymerase chain reaction for genetic counselling in Chinese haemophilia A families

Abstract

Linkage analysis is a widely used strategy for genetic counselling in haemophilia A (HA) families. We attempted to develop more informative markers closely linked to factor VIII (FVIII) gene and establish a rapid multifluorescent polymerase chain reaction (PCR) method with these markers. Five extragenic (DXS15, DXS9901, G6PD, DXS1073 and DXS1108) and one intragenic (F8Civs13) markers were examined in 118 healthy individuals and 12 HA families which had been diagnosed before. Five extragenic markers were within an interval of about 1.5 Mb to FVIII gene and located on each side of the gene. The expected heterozygote rate (HR) of DXS15, DXS9901, G6PD, DXS1073, DXS1108 and F8Civs13 were 74.97%, 79.77%, 56.06%, 59.92%, 39.97% and 47.61%, while the observed HR were 88.24%, 82.35%, 21.57%, 62.75%, 35.29% and 52.94%. When six polymorphic markers were combined together, all the studied females were informative in at least one of these markers and 29.41% of them were detected informative in three markers with the highest frequency. The diagnostic rates of DXS15, DXS9901, G6PD, DXS1073, DXS1108 and F8Civs13 in 12 haemophilia families were 75.00%, 91.67%, 41.67%, 75.00%, 33.33% and 66.67% respectively. All the genetic diagnosis was consistent with the result we analysed before and no recombination was observed. Family 1 was given as an example in this study and was found to be informative in three polymorphic markers DXS15, DXS9901 and DXS1073. The patient's sister was detected the same allele as the proband, but her male fetus did not inherit the affected allele from her, which was consistent with the result of sequencing. It was demonstrated that the multifluorescent PCR method established in this study was convenient and efficient and can be applied to carrier detection and prenatal diagnosis in HA families.

Evaluation of factor VIII polymorphic short tandem repeat markers in linkage analysis for carrier diagnosis of hemophilia A

“…Additional informative intragenic markers, such as STR increase the accuracy of analysis (33). Furthermore, Dai et al (34) observed the HR of 21.57% for G6PD and 35.29% for DXS1108 in Chinese population, which was consistent with previous report (35).…”

Improved criterion‐referenced assessment in indirect tracking of haemophilia A using a 0.23 cM‐resolution dense polymorphic marker set

“…We, therefore, propose this informativeness as minimum criterion‐referenced assessment in indirect carrier‐detection. For comparison, a previously reported hexaplex PCR assay consisting of F8 Int13 and the extragenic markers DXS15, DXS9901, IKBKGInt1, DXS1073 and DXS1108 [13], and comprising a 2.73 cM interval (i.e., 12 times the expected recombination fraction of our heptaplex assay), rendered no tested subject informative for all six markers. Thus, the heptaplex here reported is endowed with superior precision, mainly due to the small recombination fraction and high‐cumulative heterozygosity for the selected panel of markers.…”

Duchenne Muscular Dystrophy in a Female Patient with a Karyotype of 46,X,i(X)(q10)

“…The primer sequences of these markers were published in Leiden Muscular Dystrophy Pages (http://www.dmd.nl). B, The three loci in the factor VIII gene of Xq28 analysis (Lalloz et al 1991(Lalloz et al , 1994Fang et al 2006), showing that the patient has the same haplotype as her father. Square= male, Circle =female, fully shaded = affected.…”