Detection of single nucleotide polymorphisms in the mannose-binding lectin gene using minor groove binder-DNA probes.
Structural point mutations in exon 1 at codons 52, 54 and 57 and a promotor polymorphism at -221 bp of the mannose-binding lectin (MBL) gene are associated with increased susceptibility to various infectious diseases. We developed a genotyping method based on the 5' nuclease (TaqMan) assay in combination with the use of minor-groove-binder (MGB) probes in screening for these mutations/polymorphisms. In contrast to conventional probes, MGB probes have a short length and can be used for detection of mutations that are in close proximity to each other, as is the case for the structural mutations in exon 1 of the MBL gene. Results obtained with the 5' nuclease assay using MGB probes were identical with results obtained with classical techniques such as restriction fragment length polymorphism, allele-specific PCR, and sequencing. In conclusion, the 5' nuclease assay using MGB probes is useful for large-scale screening of point mutations/polymorphisms, even when they are in close proximity.
Van Hoeyveld
Erna
E
Department of Laboratory Medicine, Immunology, Gasthuisberg University Hospital, Herestraat 49, B-3000 Leuven, Belgium. erna.vanhoeyveld@uz.kuleuven.ac.be
Houtmeyers
Frans
F
Massonet
Caroline
C
Moens
Leen
L
Van Ranst
Marc
M
Blanckaert
N
N
Bossuyt
Xavier
X
D017931:DNA Primers; D015342:DNA Probes; D005838:Genotype; D006801:Humans; D037601:Mannose-Binding Lectin / Q000235:genetics*; D015336:Molecular Probe Techniques; D017354:Point Mutation; D016133:Polymerase Chain Reaction / Q000379:methods; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D012680:Sensitivity and Specificity; D019914:Taq Polymerase
D017931:DNA Primers; D015342:DNA Probes; D037601:Mannose-Binding Lectin; D019914:Taq Polymerase