HLA-DR typing using DNA amplification by the polymerase chain reaction and sequential hybridization to sequence-specific oligonucleotide probes.
A series of sequence-specific oligonucleotides (SSOs) have been used to type alleles at the HLA-DRB1 locus. Genomic DNA was amplified to high copy number by the polymerase chain reaction (PCR) and hybridizations of the dot-blotted, amplified DNA to a series of 14 SSOs enabled the identification of the major specificities DR1-DRw14. Certain alleles (DR3 and DR4) could be rapidly and accurately identified by running the products of allele-specific amplification of genomic DNA on agarose gels. This approach facilitated the typing of serological specificities such as subtypes of DR3 (DRw17 and DRw18) as well as alleles previously detected by the mixed lymphocyte reaction including subtypes of DR4 (Dw4, Dw10, Dw13, Dw14, and Dw15). The HLA-DR types obtained by SSO probing conformed to rules of Mendelian inheritance when they were applied to a series of 75 families. A full DR type could be obtained from many individuals simultaneously without needing to separate or store viable lymphocytes. Thus, this technique may have considerable implications for the analysis of disease associations with HLA class II alleles, particularly in circumstances where facilities for the initial preparation and storage of the samples may be limited.
Wordsworth
B P
BP
Institute of Molecular Medicine, John Radcliffe Hospital, Headington, Oxford, England.
Allsopp
C E
CE
Young
R P
RP
Bell
J I
JI
D000483:Alleles; D001483:Base Sequence; D005784:Gene Amplification; D006684:HLA-DR Antigens / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction
D006684:HLA-DR Antigens; D015345:Oligonucleotide Probes