HLA-B40, B18, B27, and B37 allele discrimination using group-specific amplification and SSCP method.
We developed a system for discriminating HLA-B40, B18, B27, and B37 alleles using a two-step PCR method followed by SSCP analysis. Fragments (0.8 kb) including exon 2, intron 2, and exon 3 were amplified in the first PCR. We used two sets of primers, one specific for HLA-B60-related alleles and the other specific for HLA-B61-related, B18, B27, and B37 alleles. No amplifications of other class I genes or pseudogenes were observed. In the second PCR, exon 2 and exon 3 were amplified separately, using diluents of the first PCR products as templates. HLA-B61-related, B18, B27, B37, and B60-related alleles were clearly discriminated in the SSCP analysis of the second PCR products. In a population study in which B61 alleles were analyzed, B*4003 was detected in two Japanese individuals in addition to two B61 alleles previously reported to occur in Japanese, B*4002 and B*4006. The relative frequencies of B*4002, B*4006, and B*4003 in Japanese were 58, 35, and 6%, respectively. The individuals having B*4003 are the first non-South Americans in whom this allele has been detected. The SSCP banding patterns of 18 HLA-B60-positive Japanese population samples were identical to those of a B*40012 sample for both exon 2 and exon 3. We also demonstrated that the B37 allele occurring in some Japanese is B*3701.
Bannai
M
M
Department of Research, Japanese Red Cross Tokyo Metropolitan Blood Center, Japan.
Tokunaga
K
K
Lin
L
L
Ogawa
A
A
Fujisawa
K
K
Juji
T
T
D000483:Alleles; D001483:Base Sequence; D003937:Diagnosis, Differential; D015235:HLA-B Antigens / Q000032:analysis*; D059911:HLA-B18 Antigen; D015796:HLA-B27 Antigen / Q000032:analysis*; D059912:HLA-B37 Antigen; D059947:HLA-B40 Antigen; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic / Q000235:genetics*; D018807:Polymorphism, Single-Stranded Conformational
D015235:HLA-B Antigens; D059911:HLA-B18 Antigen; D015796:HLA-B27 Antigen; D059912:HLA-B37 Antigen; D059947:HLA-B40 Antigen