Alpha-1-protease inhibitor phenotypes and serum concentrations in Thailand

Summary Alpha-1-protease inhibitor (A1PI) Pi phenotypes were determined in 429 individuals from Bangkok and 423 from the rural area of the northeast of Thailand by isoelectric focusing on thin-layer polyacrylamide gel. Variation was more marked in the Bangkok population. In the rural area, with an exclusively northeastern Thai population the Pi-alleles M, S and Z were detected. A1PI serum concentrations measured by electroimmunoassay were slightly higher in Thai than the values reported from European countries; children with Pi_M from rural areas had slightly higher A1PI serum concentrations than Pi_M children from Bangkok.     

New phenotypes of serum α 1-antitrypsin in Japanese detected by gel slab isoelectric focusing

Summary The phenotype distribution and gene frequencies of serum ₁-antitrypsin in 856 healthy blood donors in Tokyo were examined by gel slab isoelectric focusing (pH 4–6). The allele of the common subtype variant Pi M₂ was present with a frequency of 0.1099 in Japanese. A study of 23 twin pairs and their parents was in agreement with the hypothesis of autosomal codominant inheritance of Pi M subtypes. Other rare variant alleles, Pi M^F, Pi M^S, Pi M^N, Pi M^V, Pi M^X, Pi M^Z were found in very low frequencies.The total concentration of serum ₁-antitrypsin was compared among three different phenotypic groups (M₁, M₁M₂, M₂). Statistically significant quantitative differences were found among these three groups (P<0.01).     

PiT: A new allele in the alpha1-antitrypsin system

Summary In a genetic investigation of the population in Hessen, Germany, we found a family with a new, rare allele in the Pi system (₁-antitrypsin). According to electrophoretic analysis and isoelectric focusing patterns, it is designated Pi ^T. A pedigree study suggests autosomal codominant inheritance. The serum concentration of six heterozygous carriers of this allele (phenotype M1T or M2T) revealed normal ₁-antitrypsin levels.     

Beta-Galactosidase immunoassay for the measurement of cyclic AMP

The previously described method for phenotyping of alpha₁-antitrypsin (alpha₁-protease inhibitor, Pi) that utilizes separator isoelectric focusing on thin-layer agarose gel (A. R. Qureshi and H. H. Punnett, in Electrophoresis '81, 3rd International Conference on Electrophoresis, pp. 83–87 (1981)) has been improved to give a better resolution of Pi pattern. A shallow pH gradient in the region of the isoelectric point of Pi pattern was obtained by the use of N-(2-acetamido)-2 aminoethanesulfonic acid (1%) and serine (0.8%). The present technique can resolve the Pi alleles. The patterns of Pi phenotypes were found to be similar to those observed on acrylamide gels. The method is fast, reliable, and reproducible.     

α1-antitrypsin (Pi) phenotypes in a village population from the Gambia,West Africa

Summary A total of 701 individuals from a village in The Gambia, West Africa were tested for serum ₁ ^- antitrypsin phenotypes by isoelectric focusing in thinlayer polyacrylamide gels (pH 4-5). A new variant allele, Pi ^GAM , was discovered at a polymorphic frequency (0.0642), and the inheritance of the variant phenotype was confirmed by family studies. The variant was not found to be associated with any decrease in serum ₁ ^- concentration. The only other allele found within this population was the common allele Pi ^M (0.9358).     

Pi M4: An additional Pi M subtype

Summary The authors studied Pi polymorphism using the Separator isofocusing method with slight modification. A new Pi allele was observed. Family pedigrees confirmed co-dominant inheritance with other Pi alleles. According to the electrophoretic mobility of its isoprotein bands, and to its frequency (0.04) this new allele is considered as a fourth Pi ^M subtype: Pi ^M4.     

Identification of Two Blast Resistance Genes in a Rice Variety, Digu

Blast, caused by Magnaporthe grisea is one of most serious diseases of rice worldwide. A Chinese local rice variety, Digu, with durable blast resistance, is one of the important resources for rice breeding for resistance to blast (M. grisea) in China. The objectives of the current study were to assess the identity of the resistance genes in Digu and to determine the chromosomal location by molecular marker tagging. Two susceptible varieties to blast, Lijiangxintuanheigu (LTH) and Jiangnanxiangnuo (JNXN), a number of different varieties, each containing one blast resistance gene, Pik^s, Pia, Pik, Pi‐b, Pi‐k^p, Pi‐ta², Pi‐ta, Pi‐z, Pi‐i, Pi‐k^m, Pi‐z^t, Pi‐t and Pi‐11, and the progeny populations from the crosses between Digu and each of these varieties were analysed with Chinese blast isolates. We found that the resistance of Digu to each of the two Chinese blast isolates, ZB13 and ZB15, were controlled by two single dominant genes, separately. The two genes are different from the known blast resistance genes and, therefore, designated as Pi‐d(t)1 and Pi‐d(t)2. By using bulked segregation method and molecular marker analysis in corresponding F₂ populations, Pi‐d(t)1 was located on chromosome 2 with a distance of 1.2 and 10.6 cM to restriction fragment length polymorphism (RFLP) markers G1314A and G45, respectively. And Pi‐d(t)2 was located on chromosome 6 with a distance of 3.2 and 3.4 cM to simple sequence repeat markers RM527 and RM3, respectively. We also developed a novel strategy of resistance gene analogue (RGA) assay with uneven polymerase chain reaction (PCR) to further tag the two genes and successfully identified two RGA markers, SPO01 and SPO03, which were co‐segregated toPi‐d(t)1 and Pi‐d(t)2, respectively, in their corresponding F₂ populations. These results provide essential information for further utilization of the Digu's blast resistance genes in rice disease resistance breeding and positional cloning of these genes.     

Determination of isoelectric points in thin-layer isoelectric focusing: The importance of attaining the steady state and the role of CO2 interference

Isoelectric points differing by 1 to 2 pH units are measured for horseradish peroxidase and lactoperoxidase depending upon the technique of isoelectric focusing, namely, the density gradient technique or systems stabilized by either granulated (Sephadex, Bio-Gel) or compact polyacrylamide gels. Conditions standardized for the determination of pI values of selected pH marker proteins proved inadequate for the predominant isoenzyme of horseradish peroxidase which requires an excessively long focusing time to attain the steady state. Carbon dioxide interferes with the determination of pI values >8.2 to 8.3. Thin-layer isoelectric focusing in a CO₂-free atmosphere followed by pH measurements also in a CO₂-free atmosphere, yields for alkaline marker proteins and the predominant peroxidase isoenzyme, pI values in excellent agreement with these found by the density gradient technique. The isoionic point of the predominant peroxidase isoenzyme determined by ion exchange desalting is identical with the isoelectric point found by density gradient and thin-layer isoelectric focusing in a CO₂-free atmosphere.     

An excess of the Pi Sallele in dizygotic twins and their mothers

Summary A significant excess of the ₁ (protease inhibitor) Pi ^S allele has been found in 147 pairs of dizygotic (DZ) twins, but frequencies in 170 pairs of monozygotic (MZ) twins do not differ from those in a sample of 1007 blood donors. In 51 mothers of DZ twins the frequency of the Pi ^S allele was double than in the same sample of donors, but there was no corresponding increase in the fathers of DZ twins nor in the parents of MZ twins. In an independent sample of 66 mothers of twins of unknown zygosity, there was also a significant excess of Pi ^M Pi^S and Pi^M Pi^Z phenotypes, and this was particularly marked in the subsample of mothers of opposite-sex twin pairs. We speculate that lowered protease inhibitor levels in women carrying the Pi ^S allele may enhance sperm migration, increase the probability of multiple ovulation, or both.     

Polymorphism of alpha-1 antitrypsin in dogs

• 1.1. A genetically determined polymorphism of alpha-1 antitrypsin is demonstrated in dog serum by isoelectric focusing in a pH range of 3.5–5.0, followed by direct immunoblotting using a specific antiserum.
• 2.2. Alpha 1 antitrypsin focuses as two major bands at isoelectric points of 4.60 and 4.64 or 4.67 and 4.7 in presumed homozygous animals. Heterozygotes show both sets of bands.
• 3.3. The results of seven crosses with 33 offspring are best explained by two codominant alleles, Pi^M and Pi^S at a single locus designated as Pi for proteinase inhibitor.
• 4.4. The concentration of alpha-1 antitrypsin in serum of healthy dogs was 2.65 ± 0.42 mg/ml and 2.19 ± 0.38 mg/ml in females andv males respectively.
• 5.5. The higher concentration in female dogs suggests that estrogens may influence the serum level of alpha-1 antitrypsin.

     

Polymorphism analysis of genomic regions associated with broad-spectrum effective blast resistance genes for marker development in rice

Cultivated European rice germplasm is generally characterized by moderate to high sensitivity to blast, and blast resistance is therefore one of the most important traits to improve in rice breeding. We collected a panel of 25 rice genotypes containing 13 broad range rice resistance genes that are commonly used in breeding programs around the world: Pi1, Pi2, Pi5, Pi7, Pi9, Pi33, Pib, Pik, Pik-p, Pita, Pita ² , Piz and Piz-t. The efficiency of the selected Pi genes towards Italian blast pathotypes was tested via artificial inoculation and under natural field infection conditions. To characterize haplotypes present in the chromosomal regions of the blast resistance genes, a polymorphism search was conducted in the sequence regions adjacent to the blast resistance by examining DNA from the Pi gene donors with a panel of 5–7 potential receivers (cultivated European rice genotypes). Seven InDel and 8 presence/absence polymorphisms were directly detected by gel analysis after DNA amplification, while sequencing of 12.870 bp through 32 loci in different genotypes revealed 85 SNP (one SNP every 151 bp). Seven SSRs were additionally tested revealing 5 polymorphic markers between donors and receivers. Polymorphisms were used to develop 35 PCR-based molecular markers suitable for introgressing of Pi genes into a set of the European rice germplasm. For this last purpose, allelic molecular marker variation was evaluated within a representative collection of about 95 rice genotypes. Polymorphic combinations allowing introgression of the broad spectrum resistance genes into a susceptible genetic background have been identified, thus confirming the potential of the identified markers for molecular-assisted breeding.     

中国广东人红细胞PGM_1亚型遗传性多态现象

采用薄层聚丙烯酰胺凝胶等电聚焦技术,调查了中国(广东)406名无亲缘关系的正常人红细胞磷酸葡萄糖变位酶-1(PGM_1)亚型的遗传多态性。除了常见的10种亚型外,还发现了由一个新的变异型等位基因和常见的4个等位基因杂合产生的9例变异型。PGM_1位点的等位基因频率PGM_1~(1+)、PGM_1~(1-)、PGM_1~(2+)、PGM-1~(2-)和PGM_1~(V丰)(变异型等位基因)分别为0.5973、0.1256、0.1724、0.0936和0.0111;群体处于Hardy-Weinberg式平衡状态。变异型等位基因以多态频率出现,可能成为该群体的一个重要的遗传性特征。     

Evidence for two additional common alleles at the PGM1 locus (Phosphoglucomutase-E.C.:2.7.5.1)

Summary Lysates of erythrocytes, leukocytes, lymphocytes, and extracts of sperms were investigated for the PGM₁ isozymes by three techniques: starch gel electrophoresis, high voltage thin-layer agarose gel electrophoresis, and thinlayer isoelectric focusing on polyacrylamide gel. On starch, only the well known common phenotypes 1, 2-1, and 2 were demonstrable. On agarose, different distances of the two main cathodal bands (a, b) among the phenotypes 2-1 were noted. Furthermore, on agarose, some types considered as homozygous on starch gel had a single, sharp banded pattern, while others were broad and blurred. Optimal separation was achieved by isoelectric focusing on polyacrylamide gel. In 291 leukolysates, 10 different phenotypes were identified. These are considered as gene products of 4 different common alleles at the PGM₁ locus as suggested by preliminary family investigations. In a random population from Hessen these four alleles, had the following frequencies: PGM ₁ ^a1 0.6186, PGM ₁ ^a2 0.1718, PGM ₁ ^a3 0.1426, and PGM ₁ ^a4 0.067. The preliminary designation a1, a2, a3 and a4 was chosen as the assumed polymorphism was demonstrated on acrylamide and agarose. The sum of the frequencies PGM ₁ ^a1 and PGM ₁ ^a3 (the gene products of which have apparently the same electrophoretic mobility on starch) is similar to the frequency of the old PGM ₁ ¹ allele (0.757) in Caucasoids, PGM ₁ ^a2 and PGM ₁ ^a4 have a frequency of 0.2388 corresponding with the frequency of the old allele PGM ₁ ² .     

Group-specific component (Gc) subtypes in the Chinese population of Hong Kong

Summary The distribution of Gc subtypes in a sample of the Chinese population of Hong Kong was studied using isoelectric focusing followed by immunofixation. A sensitive modification of this technique is described. Nine distinct phenotypes were observed which appear to result from the three common alleles Gc ^IF, Gc ^IS, and Gc ², which are found in most populations. The respective gene frequencies were 0.494, 0.258, and 0.247. In addition, two rare phenotypes were observed which appear to be due to a rare allele tentatively identified as Gc ^2G2.     

Placental alkaline phosphatase types and subtypes determined by agarose gel electrophoresis and separator isoelectric focusing

Summary Two techniques for phenotyping the human placental alkaline phosphatase system were developed: high-voltage agarose-gel electrophoresis and thin-layer separator isoelectric focusing on agarose. These methods enabled a more rapid and sensitive phenotyping of all common phenotypes than the traditionally employed starch-gel electrophoresis. An extended polymorphism of placental alkaline phosphatase was revealed by isoelectric focusing. The existence of two suballeles of Pl¹ allele and two suballeles of Pl² allele was postulated.     

Polymorphism of α1-antitrypsin in a Portuguese population

Summary Serum Pi phenotypes were studied in 219 samples. The MM phenotype was the most common as in all other populations. The frequencies of Pi^S and Pi^Z were high considering other populations.Pi^F was not detected.This investigation was supported by a grant from Instituto de Alta Cultura (Project LMC.-10).     

Isoelectric focusing studies of human red cell esterase D: Evidence for polymorphic occurrence of a new allele EsD 7 in Japanese

Summary The isoelectric focusing study of esterase D in Japanese revealed evidence of a new polymorphic allele (EsD ⁷) which is difficult to find by conventional starch gel electrophoresis only. A comparison with the occurrence of a subdivision of EsD ² in Caucasians (EsD ⁵) suggests a remarkable difference in allele distribution of esterase D among races. Quantitative analysis showed a relatively low value of enzyme activity for this new allele. It is therefore emphasized that in addition to conventional electrophoresis, enzyme assay and further detection by isoelectric focusing are essential in analyzing the esterase D system.     

The equine protease inhibitory system (Pi): Abnormal expressions of PiF,PiL, and PiS

Three cases of abnormal expression of the equine protease inhibitory alleles, Pi F, L, and S₁, were observed following the examination of 30,000 plasma samples by one-dimensional acid (pH 4.6) polyacrylamide gel electrophoresis. Characterization of the abnormal proteins in terms of isoelectric point, molecular mass, inhibitory spectra, and sialic acid content was performed using one- and two-dimensional electrophoretic techniques. The Pi F and S₁ abnormalities were postulated to be the result of amino acid substitutions causing alterations in the processing of the carbohydrate side chains. No explanation could be offered for the Pi L abnormality other than a charge shift mutation. Abnormal types, F*, L*, and S*₁ behaved as alleles but the distribution of L* in offspring from one stallion (present in only 6 of 83 offspring) differed significantly from expectation.This work was supported by a grant from the Australian Stud Book, Alison Road, Randwick, N.S.W. 2031.     

Pi Ecincinnati: A new alpha1-antitrypsin allele in three negro families

Summary Serum specimens of three unrelated black males had an unusual alpha-1-antitrypsin phenotype, designated Pi Ecincinnati because of its electrophoretic mobility. Family studies indicated that the new phenotype was the expression of an alpha-1-antitrypsin allele, labeled Pi ^Ecincinnati     

Alpha-1-antitrypsin (Pi) types in Korean and Chinese populations

Summary Alpha-1-antitrypsin phenotypes were determined by isoelectric focusing in 270 Koreans and 52 Chinese. The frequencies of the major alleles were the following, numbers for the Chinese sample are in parentheses: Pi^M1: 0.65 (0.66), Pi^M2: 0.22 (0.25), Pi^M3: 0.06 (0.09). Other alleles, including Pi^Z were present in low frequencies. The Koreans appear to be quite similar to the Chinese in this system.