Hiroaki Nishimukai

Genotyping of the ABO blood group system: analysis of nucleotide position 802 by PCR-RFLP and the distribution of ABO genotypes in a German population

Genotypes of the ABO blood group system were studied by PCR-RFLP analysis of the eight polymorphic nucleotide positions (ups) 261, 467, 526, 646, 703, 796, 802 and 803 of the cDNA from A transferase. In 169 unrelated German individuals, 17 genotypes were found and the calculated allele frequencies of A(Pro), A(Leu), B, O(T), O(A) and O2 were 0.2130, 0.0770, 0.0473, 0.4260, 0.2160 and 0.0207, respectively. These frequency data may provide useful additional information for disputed paternity and stain testing. A variant O allele, O2, was fout at a polymorphic frequency. As the nucleotide (np 261) of the O2 allele is the same as that of A and B alleles, the analysis of at least three nucleotide positions, i.e. ups 261, 526 and 802, is necessary to avoid mistyping of the ABO genotype.

Genotyping of ABO blood groups by PCR and RFLP analysis of 5 nucleotide positions

The genotyping of ABO blood groups was performed using the polymerase chain reaction (PCR) method. The 4 DNA fragments containing the nucleotide position 261, 526, 703 and 796 of cDNA from A-transferase were amplified by PCR, and the amplified DNA subjected to restriction fragment length polymorphism (RFLP) analysis. The different nucleotide at position 803 was clearly distinguished by electrophoresis of the PCR products amplified with allele-specific primers. By analyzing the electrophoresis patterns, ABO genotyping was conclusively accomplished. The frequencies of ABO genotypes found in Japanese blood donors with A and B phenotypes were as follows: in the phenotype A group, AA =19.8 % and AO = 80.2%; and in the phenotype B group, BB =12.8% and BO=87.2%.ZusammenfassungDie Bestimmung der ABO-Blutgruppen wurde mittels Polymerase-Kettenreaktion (PCR) durchgeführt. Die vier DNA-Fragmente, die Nukleotidpositionen 261, 526, 703 und 796 von der cDNA der A-Transferase enthielten, wurden mittels PCR amplifiziert. Die amplifizierte DNA wurde einer Restriktionsfragmentl ängen-Analyse (RFLP) unterzogen. Nach Amplifikation mit allelspezifischen Primern konnte der Nukleotidunterschied an Position 803 durch Elektrophorese der PCR-Produkte klar getrennt werden. Die Bestimmung der ABO-Genotypen war durch die Analyse der eletrophoretischen Muster eindeutig durchführbar. Folgende Frequenzen der ABO-Genotypen von japanischen Blutspendern wurden für die Phänotypen A und B gefunden: In der Phänotypgruppe A, AA =19,8% und AO=80,2%, Phänotypgruppe B, BB=12,8% und BO= 87,2%.

Distribution of abo genotypes and allele frequencies in a korean population

SummaryThe genotypes of the ABO blood group system were investigated in Korean living in Kangwon-Do area by PCR-RFLP analysis of the seven polymorphic nucleotide positions 261, 467, 526, 646, 703, 796 and 803 of the cDNA from A1 transferase. In 253 unrelated Korean individuals, 15 genotypes were found and the allele frequencies of A(Pro), A(Leu), B, O(T) and O(A) were 0.022, 0.209, 0.209, 0.360 and 0.200, respectively, with no deviation from Hardy-Weinberg expectations (Χ 2=2.145, d.f.=6, 0.90<p<0.95). As for the distribution of allele frequencies, a significant difference was noticed between the Korean and a Japanese (Χ 2=30.87, d.f.=4, p<0.001) and a German (Χ 2=127.76, d.f.=4, p<0.001) populations.

Immunogenetics of early-onset insulin-dependent diabetes mellitus among the Japanese: HLA, Gm, BF, GLO, and organ-specific autoantibodies — the J.D.S. study

The Japan Diabetes Society (JDS) conducted a multicenter study on the immunogenetics of early-onset insulin-dependent diabetes mellitus (IDDM) of the Japanese. Human leukocyte antigen (HLA), properdin factor B (BF), immunoglobulin heavy-chain complex (Gm), and glyoxalase of erythrocytes (GLO) were typed, and organ-specific autoantibodies, including islet cell antibody (ICA), were assayed in 159 Japanese IDDM patients and their family members and in 258 healthy Japanese controls. The HLA-DRw9 phenotype and HLA-Bw61/DRw9 haplotype were significantly increased among the patients with autoantibodies other than ICA but with no autoimmune diseases (RR = 5.84, cP less than 0.001; and RR = 7.45, P less than 0.001), whereas the HLA-DR4 phenotype and HLA-Bw54/DR4 haplotype were significantly increased in those without either the autoantibodies or autoimmune diseases (RR = 2.64, cP less than 0.001; and RR = 4.55, P less than 0.001). The HLA-DR4 phenotype was significantly increased in the patients with autoimmune thyroid diseases (RR = 6.21, cP less than 0.05). In all groups of patients, the HLA-DR2 phenotype was significantly decreased, and the relative risk of the HLA-DRw9/DR4 genotype was highest among all HLA-DR genotypes. No significant association was found between HLA type and the duration or incidence of ICA. Gm types of g and gft were significantly increased in the patients with the autoantibodies (RR = 2.11, P less than 0.05; and RR = 34.11, P less than 0.05), whereas the BF-F phenotype was significantly decreased in the patients either with or without autoantibodies (RR = 0.43, P less than 0.05; and RR = 0.46, P less than 0.05). There was no association between IDDM and GLO type. These data indicate that immunogenetic bases underlying IDDM of the Japanese are heterogeneous, as are those in Caucasians.

Distribution of OCA2∗481Thr and OCA2∗615Arg, associated with hypopigmentation, in several additional populations

Two mutants, OCA2∗481Thr (c.1441G>A, p.Ala481Thr) and OCA2∗615Arg (c.1844A>G, p.His615Arg), in the OCA2 (oculocutaneous albinism type II) gene are associated with hypopigmentation in East Asians. Here, these two alleles were studied to assess the frequencies in five different populations. In addition, the allele frequency of OCA2∗615Arg was investigated in seven populations. Among a total of 24 global populations investigated, Oroqens in Heihe showed the highest frequency for OCA2∗481Thr (0.519), and among 26 populations, Han Chinese in Changsha showed the highest frequency for OCA2∗615Arg (0.673). This study confirmed that these two East Asian-specific alleles are characteristic of northern and central-southern East Asian populations.

Genetic Polymorphism of the Seventh Component of Complement: A New Variant

Abstract. Genetic polymorphism of the seventh component of complement (C7) was studied by the methods of agarose gel isoelectric focusing and immunoblotting. Serum or plasma samples were treated with neuraminidase prior to isoelectric focusing. A cathodal variant, named C7 5, was recognized in the treated samples. Family study indicates that the C7 5 component is genetically determined. The allele frequencies calculated from 183 healthy Japanese individuals were C7*1 = 0.809, C7*2 = 0.104, C7*4 = 0.038, and C7*5 = 0.049. An association between C7 and C6 alleles was not found.

Polymorphism of Plasminogen in Healthy Individuals and Patients with Cerebral Infarction

Phenotyping of plasma plasminogen (PLG) was carried out by the method of agarose gel isoelectric focusing followed by immunofixation or immunoblotting. The allele frequencies calculated from healthy Japanese individuals (n = 795) were as follows: PLG*1 = 0.9440, PLG*2 = 0.0189, PLG*A = 0.0076, PLG*A2 = 0.0006, PLG*B = 0.0138, PLG*B2 = 0.0013, and PLG*C = 0.0138. The PLG phenotype distribution in a group of patients with cerebral infarction (n = 125) was also studied. The allele frequencies were PLG*1 = 0.960, PLG*2 = 0.016, PLG*A = 0.012, and PLG*B = 0.012. No statistically significant association was found between PLG types and cerebral infarction.

Geographical North-South Decline in DNASE1*2 in Japanese Populations

Allele frequencies for human deoxyribonuclease I (DNase I) phenotypes were determined using blood samples from about 2000 Japanese subjects living in nine prefectures, and compared with one another. DNase I phenotyping was performed principally using isoelectric focusing electrophoresis and activity staining. The DNase I system was shown to have enhanced potential for anthropologic, genetic, and clinical studies of Japanese populations. DNase I phenotypes were analyzed to evaluate the degree of genetic variation at the DNASE1 locus. Our examination of DNase I types revealed a decreasing north-to-south gradient in the DNASE1allele.

I(C3b/C4b inactivator) typing by agarose gel isoelectric focusing and immunoblotting technique

Genetic polymorphism of I (C3b/C4b inactivator) was studied by the method of agarose gel isoelectric focusing followed by an immunoblotting technique. Serum or plasma samples were pretreated with neuraminidase. The method is rapid, and gives the simple and reliable patterns of I. The allele frequencies calculated from healthy Japanese individuals living in the western part of Japan were: IF* A = 0.126 and IF*B = 0.874.

C6 Polymorphism in Japanese: Typing by Agarose Gel Isoelectric Focusing-Immunofixation

Agarose gel isoelectric focusing was used to investigate the genetic polymorphism of the sixth component of complement (C6) in Japanese. C6 patterns were visualized by the immunofixation procedure. The allele frequencies calculated from 135 individuals were as follows: C6*A = 0.467, C6*B = 0.481, C6*B2 = 0.037, and C6*B3 = 0.015. It is suggested that C6*B3 is the fourth common allele characterizing the Japanese population.