Discrimination of human HLA-DRB1 alleles by PCR-SSCP (single-strand conformation polymorphism) method.
A single-strand conformation polymorphism (PCR-SSCP) method has been adopted for discrimination of human HLA-DRB1 alleles. This method enabled the detection of DNA polymorphisms including point mutations at a variety of positions in the DNA fragments of the HLA-DRB1 gene. A total of 27 HLA-DRB1 alleles from 172 healthy donors were analysed using a combination of PCR-SSCP with group-specific amplifications. Application of a small amount of amplified and denatured DNA to non-denaturing electrophoresis followed by silver staining resulted in distinct banding patterns. Samples possessing a single allele in each amplification group showed two-band patterns which correspond to the sense and antisense strands, while heterozygotes in the same group or a mixture of two single-type samples showed four-band patterns. All of the analysed alleles were discriminated in each DRB1 group. The method described here may be somewhat complicated for routine typing of HLA-DRB1 alleles. However, it is useful in the screening of "new' alleles as well as the donor-recipient molecular matching of HLA class II genes for various purposes, e.g. selection of bone marrow transplant donors.
Bannai
M
M
Department of Research, Japanese Red Cross Tokyo Metropolitan Blood Center, Japan.
Tokunaga
K
K
Lin
L
L
Kuwata
S
S
Mazda
T
T
Amaki
I
I
Fujisawa
K
K
Juji
T
T
D000483:Alleles; D004247:DNA; D006684:HLA-DR Antigens / Q000032:analysis / Q000145:classification / Q000235:genetics*; D059811:HLA-DRB1 Chains; D006801:Humans; D016376:Oligonucleotides, Antisense; D016133:Polymerase Chain Reaction / Q000379:methods; D018807:Polymorphism, Single-Stranded Conformational
D006684:HLA-DR Antigens; D059811:HLA-DRB1 Chains; D016376:Oligonucleotides, Antisense; D004247:DNA