Linkage disequilibrium between two highly polymorphic microsatellites.

The PCR was used to amplify genomic DNA from two microsatellite (dC-dA)n.(dG-dT)n sequences found to be present in the same chromosome 5 genomic clone. Analysis of the haplotype frequencies of these two interspersed repeat sequences in individuals showed strong allelic association or linkage disequilibrium. Six alleles were found for p599 (CA)n with a PIC value of 0.71 and 8 alleles were seen for lambda 599 (CA)n with a PIC value of 0.74. The two microsatellites are separated by approximately 7 kb. Analysis of the length variations for the two microsatellites showed that they were positively correlated, a finding that has no obvious explanation. The strong linkage disequilibrium found demonstrates stability during evolution for these novel markers. Therefore they should be powerful new tools for studying genetic drift and admixture of populations. Furthermore, disequilibrium data from microsatellites can be used in the fine mapping and cloning of disease genes.     

Carbonic anhydrase isoenzymes in nine troops of Kenya baboons, Papio cynocephalus (Linnaeus 1766)

The distributions of alleles at the carbonic anhydrase I (CA I = CA B) and carbonic anhydrase II (CA II = CA C) loci in nine troops of Papio cynocephalus were determined. Two alleles were found at the CA I locus, and three at the CA II locus; the frequencies were: CA I^a = 0.856; CA I^b = 0.144; CA II^a = 0.784; CA II^b = 0.209; CA II^c = 0.007. Results of tests for Hardy-Weinberg equilibrium, homogeneity tests, and calculations of migration rates were used in support of the interpretation that migration and genetic drift may affect the distribution of alleles at the CA I locus and that selection is the process responsible for the distribution of alleles at the CA II locus.     

Antigen levels of urokinase-type plasminogen activator receptor and its gene polymorphism related to microvessel density in colorectal cancer

We determined the distribution of genotypes and frequencies of alleles of the (CA)(n) repeat polymorphism in intron 3 of the urokinase plasminogen activator receptor (uPAR) gene, uPAR antigen levels and microvessel density (MVD) in tumour and distant mucosa samples from 52 patients with colorectal cancer. The uPAR level was higher for patients with high MVD comparing to patients with lower MVD which may suggest that uPAR can be correlated with progression of colorectal cancer. The significant relationship between the high MVD and uPAR antigen level appeared to be independent of the (CA)(n) repeat polymorphism because no differences in the level of uPAR antigen between carriers of alleles were found. The received results, indicate that uPAR might be considered as a target in colorectal cancer patients' therapy.     

Four DNA polymorphisms on the short arm of the X chromosome: allele frequencies in a German and in a Turkish population

Four restriction fragment length polymorphisms, revealed by cloned arbitrary X chromosome segments (L1.28, RC8, pD2, 754) were studied in samples (50 individuals each) of a German and a Turkish population. All previously reported alleles of these polymorphisms were found in both populations, except the infrequent RC8 allele B3 (3.0 kb fragment), which was absent in both groups. The observed minor alleles were found to be rarer in the German series than in the Turkish group, but there was no conclusive evidence of essentially different allele frequencies in either population. However, the frequencies of the RC8 allele B2 (5.3 kb fragment) were differing at the 5% significance level. The allele frequencies of the four polymorphisms are presented and compared with those reported from other European regions.     

Haemochromatosis gene mutations in Finns, Swedes and Swedish Saamis

Frequencies of three different mutant haemochromatosis (HFE) alleles (282Tyr, 63Asp and 65Cys) were studied in three northern European populations, i.e. Finns, Swedes and Swedish Saamis. In Finns and Swedes the allele frequencies were within the range found in other populations from northern and western Europe. The Saamis differed from the Swedes with respect to all mutant alleles. Lower frequencies compared to Swedes were found for the 282Tyr (p = 0.0046) and 63Asp (p = 0.034) alleles, whereas the frequency of the 65Cys allele was higher (p = 0.046) in the Saamis. The total distribution of HFE alleles in Saamis showed a highly significant difference from that in Swedes (chi2 = 16.7, 3 d.f., p = 0.0008). These results further underline the genetic uniqueness of the Saamis.     

Carbonic anhydrase II in new world monkeys

Carbonic anhydrase II electrophoretic patterns were investigated in 3113 animals belonging to 12 genera and 24 species of New World primates. Polymorphism was detected in 13 species. A total of 24 different alleles was postulated to explain the variability found; the genusAotus showed the highest (eight) number of such alleles. Three genera of the family Callitrichidae (Callithrix, Saguinus, andCebuella) presented five alleles that were not found among the Cebidae. Important markers at the generic level were observed inCallicebus (CA2 ^*6 andCA2 ^*12),Cebus (CA2 ^*10, CA2^*16, andCA2 ^*21), andAotus (CA2^*3, CA2^*4, CA2^*5, CA2^*9, CA2^*15, CA2^*17, CA2^*22, CA2^*23). CA2^*13 seems to be the most common allele among the Cebidae; six genera of this family showed frequencies higher than 70% of it.     

Analysis of intragenic polymorphic markers of the CFTR gene in cystic fibrosis patients and health donors from Bashkorostan

The differences in the polymorphic allele frequency distribution patterns of the biallelic (M470 and TUB20) and microsatellite (IVS6aGATT, IVS8CA, and IVS17CA) markers within the CFTR gene between normal and delF508 chromosomes have been established. For most of the marker loci similar distribution of the allele frequencies on normal and mutant chromosomes without delF508 was demonstrated. Certain polymorphic alleles displayed substantial linkage disequilibrium with the delF508 mutation. Analysis of the IVS6aGATT-IVS8CA-M470-IVS17CA-TUB20 haplotypes association on normal and mutant chromosomes provided identification of the delF508 ancestral haplotype. It was suggested that delF508 mutant chromosomes were introduced into the modern Bashkir gene pool as a result of Slavic migrations from the Eastern Europe. The IVS6aGATT-IVS8CA-M470-IVS17CA-TUB20 major haplotype (77272) revealed was statistically significantly most frequently found on the mutant chromosomes without the delF508 mutation. This finding suggests that the Bashkir cystic fibrosis patients, mostly belonging to the Turkic-speaking families, possessed specific CF gene defect associated with the given haplotype.     

Investigation of serum protein systems in Chinese pigs

Polymorphisms of transferrin (Tf), pre-albumin (Pa), haemopexin (Hpx), ceruloplasmin (Cp) and amylase (Am) of Duroc pigs and Hunan indigenous pigs were investigated using horizontal starch gel electrophoresis. Allele frequencies of Durocs determined in 2004 were compared with frequencies presented in the paper by Baker L.N. (1968) Serum protein variation in Duroc and Hampshire pigs. Vox Sanguinis15, 154-8. The number of serum protein alleles decreased over time and allele frequencies aggregated across certain alleles, including TfB, PaA, Hpx3, CpB and AmB. Differences in allele frequencies, average heterozygosities and standard genetic distances between the Duroc pigs and Hunan indigenous pig populations were examined. The relationship between Durocs and Hunan indigenous pigs was found to be more distant than those among the three Hunan indigenous pig populations.     

C8A and C8B polymorphisms in Norwegians and Norwegian lapps

C8 inheritance patterns in 364 mother-child pairs formed the basis for evaluation of the existence of silent alleles (null alleles) in the genes determining the two known polymorphic C8 systems. While evidence for such alleles was not found in C8A (alpha-gamma complex), two observations of null allele segregation in C8B (beta chain) indicate a C8BQ*0 allele frequency of about 0.07. Two population samples comprising 150 Lappish and 1,264 non-Lappish Norwegians were examined for phenotype distributions in C8A and C8B. The phenotype distributions were mainly in accordance with the expected Hardy-Weinberg distribution. The results for C8A indicated simple, codominant inheritance of two frequent and several rare alleles. Allele frequencies were similar in the two populations. The C8A B gene frequency in Norwegians was significantly lower than that in FRG and higher than that in Negroes. C8B allele frequencies were also calculated from gene counts in the population material, but with due corrections for the C8BQ*0 frequency observed in the mother-child material.     

广州地区106例汉族人群MICA和MICB微卫星基因座 单体型研究

利用聚合酶链反应和荧光（6－FAM）自动化检测技术对广东地区汉族106例无亲缘关系样本进行MICA基因外显子5和MICB基因内含子1微卫星基因座多态性及其单体型分布调查。根据群体资料估算两者间的单体型频率、连锁不平衡参数、相对连锁不平衡参数。结果显示,广州地区汉族人群MICA和MICB微卫星基因座基因型分布符合Hardy-Weinberg平衡法则,共检出MICA微卫星基因座 5个等位基因, MICB微卫星基因座14个等位基因。其中MICA A5基因频率最高（0.2877）,A4基因频率最低（0.1321）。MICB CA14等位基因频率最高（0.3255）,CA19、CA28等位基因频率最低（0.0047）,未检出CA27。21种MICA－MICB单体型频率大于1%(连锁不平衡参数>0), 其中单体型A5－CA14 (16.73%), A5.1－CA18 (8.75%), A4－CA26(3.76%),A9－CA15(3.66%)和A6－CA21(2.61%)为强连锁常见单体型（χ2>3.84, P<0.05）。广州地区汉族人群MICA和MICB微卫星基因座多态性和单体型分布有其自身特点,MICA和MICB微卫星基因座适合做为遗传标志,用于人类学、遗传疾病基因连锁分析、法医学亲子鉴定和个体识别等研究领域。Abstract: This study is to investigate genetic polymorphisms and haplotypes of microsatellite locus in the exon 5 of the MICA gene and intron 1 of the MICB gene based on 106 samples of Guangzhou Han Population by polymerase chain reaction and fluorescent technique (6-FAM). The corresponding haplotype frequencies, linkage disequilibria values and relative linkage disequilibria values were estimated based on population data. The results show that the genotype distributions of MICA and MICB microsatellite meet Hardy-Weinberg equilibrium in Guangdong Han population. In total, 5 alleles of MICA microsatellite locus and 14 alleles of MICB microsatellite locus were observed. MICA A5 was the most common allele (0.2877), whereas A4 was the least popular one (0.1321). MICB CA14 was the most common allele (0.3255), and CA19 and CA28 were the least popular ones (0.0047). CA27 was not observed. Twenty-one kinds of MICA-MICB haplotypes occurred at frequencies of more than 1% (linkage disequilibria value>0). The common MICA-MICB haplotypes were A5-CA14（16.73％）, A5.1- CA18 (8.75%), A4- CA26(3.76%),A9-CA15(3.66%) and A6-CA21(2.61%)（χ2>3.84, P<0.05）, and they were strong linkage disequilibria. The polymorphisms and haplotypes distributions of MICA and MICB microsatellite locus in Guangzhou Han population have their own genetic characteristics. The microsatellite locus of the exon5 of the MICA gene and intron 1 of the MICB gene could be used as the genetic markers in the studies of anthropology, linkage analysis of genetic disease genes, individual identification and paternity test in forensic medicine.     

猪PAC克隆的微卫星DNA分离研究

利用(CA)8核心序列,设计锚定引物,采用直接测序法,从单个猪PAC克隆中分离到一个新微卫星DNA。根据该微卫星DNA的侧翼序列,设计了专一引物,在8个猪种40个个体中检测到3个等位基因,片段长度分别为305 bp、307 bp和309 bp。3种等位基因纯合子个体的PCR产物序列分析表明,这3种等位基因分别有12、13和14次CA双核苷酸重复。 Abstract:A novel microsatellite DNA was isolated from a single porcine PAC clone by sequencing the PAC clone directly with (CA)n repeat motif anchored primer.The specific primer pairs flanking the (CA)n repeat region were used to amplify the genomic DNA of 40 individuals from 8 pig breeds,which detected three alleles with the fragment length of 305 bp,307 bp and 309 bp.The PCR product sequencing results of homozygous animals representing three alleles revealed that those three alleles contained 12,13 and 14 CA dinucleotide repeats respectively.     

Susceptibility to relapsing-progressive multiple sclerosis is associated with inheritance of genes linked to the variable region of the TcR beta locus: use of affected family-based controls.

We tested the hypothesis that susceptibility to relapsing-progressive (RP) (but not to relapsing-remitting [RR]) multiple sclerosis (MS) is associated with a gene linked to the TcR beta-chain variable region delimited by the Vbeta8-BamHI and Vbeta11-BamHI RFLP alleles in DRw15+ MS patients, using a contingency-table test of patient data and affected family-based controls. Control alleles and haplotypes were composed of parental marker alleles and haplotypes not transmitted to the affected child, in 90 simplex and 31 multiplex families from British Columbia. A total of 6,164 alleles at 11 loci were segregated through families of probands with RP MS or RR MS. The Vbeta8-Vbeta11 subhaplotype frequencies in the DRw15+ RP MS (but not RR MS) patients differed from control frequencies, because of an increase of the 2-1 subhaplotype (P=.02). Vbeta8-BamHI and Vbeta11-BamHI allele frequencies (P=.05 and .009, respectively) in the DRw15+ RP MS (but not RR MS) patients differed from control frequencies. The Vbeta1-Vbeta8 subhaplotype frequencies in the DRw15- RP MS (but not RR MS) patients differed from control frequencies (P=.03), with a significantly increased frequency of the 1-1 subhaplotype (P=.01; RR=7.1) in RP MS versus RR MS patients. Susceptibility to RP MS is associated both with a recessive inheritance of a gene linked to the 3' (Vbeta11) end of the 2-1 subhaplotype defined by the Vbeta8-BamHI and Vbeta11-BamHI alleles in DRw15+ patients and with a gene, located on the 1-1 subhaplotype, defined by the Vbeta1-TaqI and Vbeta8-MspI alleles of the TcR beta-chain complex in DRw15- patients.     

HUMTH01 allele distribution in four human samples. An interpopulation analysis

The analysis of STR HUMTH01 in four populations (Galicia, SE Spain, Morocco, Cape Verde) was carried out, with up to 8 alleles being found. The frequencies of these alleles determine values of heterozigosity ranging from 0,826 detected in Cape Verde to 0,778 in Morocco. A global comparison of world populations shows clear patterns of differentiation among the main human groups, both by way of the analysis of allele distribution profiles and by the application of a Correspondence Analysis. Caucasian populations are mainly characterised by the highest frequencies in allele 9.3, Mongoloids by a very high frequency of allele 9 and Negroids show the highest frequencies for allele 7 and mainly 8. A significant negative correlation (r=−0,916, p<0,001) found in the relationships between alleles 9 and 9.3 seems to suggest the participation of non-random evolutionary forces in the population dynamics of this marker.     

Analysis and frequency of bovine lymphocyte antigen (BoLA-DRB3) alleles in Iranian Holstein cattle

The bovine lymphocyte antigen (BoLA-DRB3) gene encodes cell surface glycoproteins that initiate immune response by presenting processed antigenic peptides to CD4 T helper cells. DRB3 is the most polymorphic bovine MHC class II gene which encodes the peptide-binding groove. DRB3 gene has been extensively evaluated as a candidate marker for association with various bovine diseases and immunological traits. This study describes genetic variability in the BoLA-DRB3 in Iranian Holstein cattle. This is the first study of the DNA polymorphism of the BoLA-DRB3 gene in Iranian Holstein cattle. Hemi-nested PCR-RFLP method is used for identification the frequency of BoLA-DRB3 alleles. The BoLA-DRB3 locus is highly polymorphic in the studied herd (26 alleles). Almost 67% of the alleles were accounted for four alleles (BoLA-DRB3.2*8, *24, *11 and *16) in Iranian Holstein cattle. The DRB3.2*8 allele frequency (26.6%) was higher than the others. The frequencies of the DRB3.2*54, *37, *36, *28, *25, *14, *13, *10, *1 alleles were lower than 1%. Significant distinctions have been found between Iranian Holstein cattle and other cattle breeds studied. In Iranian Holstein cattle the alleles (BoLA-DRB3.2*22, *2 and *16) associated with a lower risk of cystic ovarian disease in Holstein cattle are found. The alleles associated with the resistance to mastitis and to bovine leukemia virus infection BoLA-DRB3.2*11 and *23 are detected with the frequencies 10.4% and 4.4%, respectively. Thus in the Iranian Holstein cows studied are found alleles which are associated with resistance to various diseases. The method of DNA-typing of animals can be used in agricultural practice for BoLA-DRB3 allele genotyping of cattle in order to reduce spreading of alleles providing susceptibility to mastitis or leukemia in cattle herds.     

A polymorphic (CA)n repeat element maps the human glucokinase gene (GCK) to chromosome 7p.

A compound imperfect dinucleotide repeat element, [CA]4TTTGT[CT]7[CA]9AA[CA]4CCACATA[CA]3, was found approximately 10 kb 3' to the human glucokinase gene (GCK) from analysis of contiguous genomic DNA obtained from a bacteriophage lambda chromosome walk. Direct human genomic sequencing revealed the source of polymorphism to be variable numbers of CT and CA repeats. Altogether six alleles that range in length from +10 to -15 nucleotides compared to the most common (Z) allele have been identified. Alleles Z, Z + 2, and Z + 4 were present in American Blacks, Pima Indians, and Caucasians, with somewhat varied frequencies among the groups. Two alleles, Z + 10 and Z - 15, appear to be unique to American Blacks, while a Z + 6 allele was observed only in the Caucasian population studied. Observed heterozygosity of the polymorphism in the CEPH reference pedigree collection is 44% and the PIC 0.44. The polymorphism is assayed by PCR amplification and resolution of 32P-end-labeled products (ranging in length from 180 to 205 bp) on denaturing polyacrylamide sequencing gels. Using the PCR assay, the human glucokinase gene was physically localized to chromosome 7 in a panel of rodent/human somatic cell lines. Genetic analysis in CEPH pedigrees placed the dinucleotide repeat element, and thereby the human glucokinase gene, on chromosome 7p between TCRG and a RFLP locus D7S57. The glucokinase dinucleotide repeat genetic marker can now be used to assess the role of the glucokinase gene in diabetes by population association studies. In addition, this repeat marker and others flanking it on chromosome 7 can be used in linkage studies with families segregating the disorder.     

Microsatellite genotyping of Chromosome 14q13.2-14q13 in the vicinity of proteasomal gene<Emphasis Type="Italic"> PSMA6</Emphasis> and association with Graves’ disease in the Latvian population

The 270-kb Chromosome 14q13.2-14q13 region harboring the proteasomal alpha subunit 6 gene PSMA6 was analyzed for polymorphism of five microsatellite repeats in cases/controls and association with Graves disease. Four novel microsatellite markers were localized to the 14q13.2 region upstream of PSMA6. Dinucleotide repeats HSMS801, HSMS702, HSMS701 were identified in two introns of the gene KIAA0391; the most upstream trinucleotide HSMS602 marker was found in an intron of the C14orf24 gene. A polymorphism study performed on the Latvian population revealed 13 and 14 alleles for HSMS801 and HSMS702, respectively, seven alleles for HSMS701, and four alleles for HSMS602. Heterozygosity analysis revealed that all the four markers obey Hardy-Weinberg distribution. The previously described HSMS006 marker, represented by 12 alleles, is localized in intron 6 of the PSMA6 gene. No significant differences were observed between patients and controls in allele distribution of the HSMS702 and HSMS701 microsatellite repeats. However, the allele frequencies of HSMS006 and HSMS801 were significantly different between Graves disease and control subjects. The 181- and 185-bp alleles of HSMS006 and the 133-, 143-, and 149-bp alleles of HSMS801 were found more often, but the 189- and 191-bp alleles of HSMS006 were much less frequent in Graves disease patients compared with the controls. An additional 174-bp allele of the HSMS602 marker, absent in healthy subjects, was found in Graves disease patients.     

Long-term response to artificial selection with multiple alleles--study by simulations

The effect of multiple alleles on long-term response to selection is examined by simulations using a pseudosampling technique to simulate the multidimensional diffusion process. The effects of alleles are independently drawn from a normal distribution and the initial frequencies of alleles are assumed either to be equal or to be drawn from a neutral equilibrium population. With these two initial gene frequency distributions we examined various properties of the selection response process for the effects of number of alleles and selection intensity. For neutral initial frequencies the effects of multiple alleles compared with two alleles are minor on the ratio of final to initial response (E(R infinity/E(R1)) and the half life of response (t0.5), but are significant on the variance of response. Under certain conditions the variance of the selection limit can even increase as selection gets stronger. For equal initial frequencies the effects of multiple alleles are, however, minor on the ratio of the variance of the selection limit to the initial genetic variance, but E(R infinity/E(R1) and t0.5 increase as the number of alleles increases. The results show that for certain statistics the effects of multiple alleles can be minimized by an appropriate transformation of parameters for given initial gene frequencies, but the effects cannot, in general, be removed by any single transformation or reparameterization of parameters.     

Novel Microsatellites from Asian Sea Bass (Lates Calcarifer) and Their Application to Broodstock Analysis

We isolated novel dinucleotide, trinucleotide, and tetranucleotide microsatellites from the genome of Asian sea bass (Lates calcarifer). Two genomic DNA libraries were established, one was enriched for (CA)n repeats, while the other for (GATA)n, (GACA)n, and (AAC)n repeats. Sixty clones containing an insert between 250 and 1000 bp in size were sequenced from each library; altogether 50 (43%) of them contained microsatellites. Forty microsatellites were characterized in 16 unrelated Asian sea bass individuals. Twenty-eight of them (70%) showed specific amplification and polymorphism. The allele number per loci varied between 2 and 20 with an average of 5.3, while expected heterozygosity ranged from 0.31 to 0.95 with an average of 0.64. At some loci allele sizes spread over a wide range (>100 bp). No significant correlation (r = 0.23, df = 31, P > 0.05) was found between the repeat number and the number of alleles. A whole broodstock containing 170 individuals was analyzed by using 8 selected polymorphic microsatellites. The average number of alleles per locus was 11.8 (range, 4–21). The expected heterozygosity ranged from 0.57 to 0.90 with an average of 0.75, while the fixation index was 0.02. Genetic similarity between individuals ranged from 0 to 0.72. Comparison of allele frequencies between the broodstock and the 24 nonrelated individuals revealed some unique alleles.     

Association of mutant alleles of serum cholinesterase with various multi-factorial and infectious diseases

Polymorphism of serum cholinesterase (SCE, acylcholinacylhydrolase, EC 3.1.1.8) for the E1 locus was studied in the groups of the patients affected with schizophrenia, peptic ulcer, hereditary erythrocytopathies, tuberculosis, thyreotoxicosis, essential hypertension and rheumatic disease. Increased frequencies of I phenotypes (E1uE1a genotype) were found among patients with peptic ulcer (12.3%), hereditary erythrocytopathies (23.2%), and UF phenotypes (E1uE1f genotype) were observed among patients with schizophrenia (2.8%) and tuberculosis (5.4%). The increased frequencies of E1a and E1f alleles in these groups of patients were, as compared to the control group, statistically significant. The value of relative risk for peptic ulcer was 3.39 in individuals of the E1uE1a genotype, those being 3.62 for schizophrenia and 6.92 for tuberculosis in individuals of the E1uE1f genotype. The nature of the other associations is discussed.     

Isolation and characterization of polymorphic microsatellites in Cocos nucifera L.

Microsatellites or simple sequence repeats (SSRs) were isolated from coconut (Cocos nucifera) and tested for polymorphism on restricted germplasm. Sequencing of 197 clones from a cv. Tagnanan Tall-enriched genomic library showed that 75% contained a microsatellite, of which 64% were dinucleotide (GA/CT, CA/GT and GC/CG), 6% were trinucleotide, and 30% were compound repeats. Of 41 primer pairs tested on Tagnanan Tall genomic DNA, 38 gave the expected size product, two amplified two loci, and another gave a multilocus pattern. On 20 coconut samples, the 38 SSRs detected 198 alleles (average: 5.2 alleles per microsatellite). Genetic diversity (D = 1 - sigma pi2) values ranged from 0.141 to 0.809. Heterozygotes were present at high frequencies among some dwarf samples. Analysis of similarity matrices based either on shared alleles at each locus (simple matching coefficient) or on allele bands across all loci (Jaccard coefficient) showed similar results. Dwarfs grouped separately from talls and showed less genetic diversity. In a wider test on 40 samples, 8 SSRs detected 64 alleles (average: eight alleles per microsatellite). These results indicate the high potential of microsatellites to detect genetic diversity in coconut germplasm.