Locked nucleic acid (LNA) probes in high-throughput genetic analysis: application to an assay for type 1 diabetes-related HLA-DQB1 alleles.
OBJECTIVE
In large-scale genetic screening, an assay that is reliable, fast and easy to perform, and straightforwardly adapted to new analytes is a necessity. We describe a one-step assay for analyzing HLA-DQB1 alleles which are associated with susceptibility to type 1 diabetes.


METHODS
The assay is based on asymmetric PCR amplification and a homogeneous hybridization method. The specificity of the probes was improved by substituting LNA (locked nucleic acid) for DNA at the critical bases.


RESULTS
The functionality of the LNA containing probes was found to be superior compared to probes consisting of DNA only. The homogeneous assay gave a correct genotyping result in 100% of the cases, which included both extracted DNA samples and blood samples dried on sample collection cards.


CONCLUSIONS
This homogeneous approach provides a simple method to define disease risk associated with HLA alleles for large-scale screening projects.
Kiviniemi
Minna
M
Department of Virology and MediCity Research Laboratory, University of Turku, Tykistökatu 6 A, 4th floor, FIN-20520 Turku, Finland. minna.kiviniemi@utu.fi
Nurmi
Jussi
J
Lövgren
Timo
T
Ilonen
Jorma
J
D001483:Base Sequence; D001800:Blood Specimen Collection / Q000379:methods; D015342:DNA Probes; D003890:Desiccation; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics*; D005063:Europium; D005470:Fluorometry / Q000379:methods; D005820:Genetic Testing / Q000379:methods*; D006683:HLA-DQ Antigens / Q000235:genetics*; D059866:HLA-DQ beta-Chains; D006801:Humans; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; D016133:Polymerase Chain Reaction / Q000379:methods*
D015342:DNA Probes; D006683:HLA-DQ Antigens; D059866:HLA-DQ beta-Chains; C069051:HLA-DQB1 antigen; D009841:Oligonucleotides; D016376:Oligonucleotides, Antisense; C477371:locked nucleic acid; D005063:Europium