Oligonucleotide ligation assay: applications to molecular diagnosis of inherited disorders

Abstract

Oligonucleotide ligation assay combined with polymerase chain reaction (PCR-OLA) is a technique which can be used for the detection of characterized sequence variations. In the present study, new PCR-OLA methods were developed for the detection of the major mutations causing infantile neuronal ceroid lipofuscinosis (INCLFin), congenital nephrotic syndrome of Finnish type (NPHS1 FinMajor and FinMinor) and medium chain acyl-CoA dehydrogenase deficiency (MCAD A985G). The prevalence of these mutations in the Finnish population was studied by analyzing blood samples collected in eastern Finland. The throughput of PCR-OLA was further enhanced by optimizing the direct use of dried blood spot (DBS) specimens for PCR. This study demonstrated that PCR-OLA is an accurate method for the detection of gene defects causing inherited disorders. With automation, PCR-OLA can be applied for routine diagnosis and for carrier screening from a large number of specimens.

Selection of a Platform for Mutation Detection

“…Ligated probes are then captured on a streptavidin-coated support and detected by alkalinephosphatase-coupled antibodies against digoxygenin. Homozygous samples generate a signal in reaction 1 or 2 only, while heterozygous samples yield a signal in both (UNIT 2.6; for a more recent application of this method, see Romppanen, 2001). ELISA-based OLA can be completely processed in microtiter plates, and the elimination of an electrophoresis step creates the possibility for high-throughput, semi-automated processing (Nickerson et al, 1990;Kwok et al, 1992).…”