Rapid and reliable genotyping of HLA-B*27 in the Chinese Han population using a duplex real-time TaqMan PCR assay.

OBJECTIVES

To develop a duplex real-time TaqMan PCR assay for genotyping HLA-B*27 in the Chinese Han population.

DESIGN AND

METHODS

A standard curve was constituted to deduce amplification efficiency, dynamic range and detection limit of the duplex real-time TaqMan PCR assay, whereas PCR-SBT (PCR with sequence-based typing) was used to evaluate the accuracy of the assay.

RESULTS

A linear standard curve for determining HLA-B*27 was obtained within the range of 10(1)-10(9) copies per reaction with the correlation coefficient of 0.99 and amplification efficiency of 98.30%. The detection limit was 3.09 copies per reaction. Complete concordance was found between the results obtained by the duplex real-time TaqMan PCR assay and PCR-SBT. Fifty-nine of the 178 genomic samples were HLA-B*27 positive and the other 119 were HLA-B*27 negative.

CONCLUSIONS

The duplex real-time TaqMan PCR approach appears to be a reliable, sensitive, rapid and high-throughput method to genotype HLA-B*27 in the Chinese Han population.