Back to Search Start Over

Allelic drop-out may occur with a primer binding site polymorphism for the commonly used RFLP assay for the -1131T>C polymorphism of the Apolipoprotein AV gene

Authors :
Hattersley Andrew T
Frayling Timothy M
Yajnik Chittaranjan S
Ellard Sian
Ward Kirsten J
Venigalla Prathyusha NS
Chandak Giriraj R
Source :
Lipids in Health and Disease, Vol 5, Iss 1, p 11 (2006)
Publication Year :
2006
Publisher :
BMC, 2006.

Abstract

Abstract Apolipoprotein AV (ApoAV) gene variant, -1131T>C, is associated with increased triglyceride concentrations in all ethnic groups studied. An MseI based RFLP analysis is the most commonly used method for genotyping this SNP. We genotyped a large cohort comprising 1185 Asian Indians and 173 UK Caucasians for -1131T>C using an ARMS-PCR based tetra-primer method. For quality control, we re-genotyped approximately 10% random samples from this cohort utilizing the MseI RFLP, which showed a 2.9% (3/102) genotyping error rate between the two methods. To investigate further, we sequenced the 900 bp region around the -1131T>C polymorphism in 25 Asian Indians and 15 UK Caucasians and found a number of polymorphisms including the -987C>T polymorphism. Further analysis of the -987C>T SNP showed a higher rare allele frequency of 0.23 in Asian Indians (n = 158) compared to 0.09 in the UK Caucasians (n = 157). This SNP is located 4 bp from the 3' end of the RFLP forward primer and is in weak linkage disequilibrium with -1131T>C variant (r2 = 0.084 and D' = 1). Repeated RFLP analysis of seven subjects heterozygous for -987C>T (seven times), showed discordant results with the sequence at -1131T>C SNP nearly one third (15/49) of the time. We conclude that presence of -987C>T polymorphism in the forward primer of the MseI RFLP assay may lead to allelic drop-out and generate unforeseen errors in genotyping the -1131T>C polymorphism. Our results also emphasise the need for careful quality control in all molecular genetic studies, particularly while transferring genotyping methods between various ethnic groups.

Details

Language :
English
ISSN :
1476511X
Volume :
5
Issue :
1
Database :
Directory of Open Access Journals
Journal :
Lipids in Health and Disease
Publication Type :
Academic Journal
Accession number :
edsdoj.7e4b504214084b7486b8081aac2d8c73
Document Type :
article
Full Text :
https://doi.org/10.1186/1476-511X-5-11