Genetic polymorphism of properdin factor B (BF) in domestic rabbit

Genetic polymorphism of plasma properdin factor B (BF) was detected in domestic rabbit, Oryctolagus cuniculus , by means of isoelectric focusing and immunoblotting. The analysis of 298 individuals, corresponding to one French and two Portuguese populations, revealed the existence of six alleles, of which BF*A, B and C were common alleles, and D, F and G were rare ones.     

A new PCR-RFLP within the domestic pigeon (Columba livia var. domestica) cytochrome b (MTCYB) gene

A total of 244 domestic pigeons (Columba livia var. domestica) were genotyped using the PCR-RFLP method. A 999 bp fragment of the MTCYB gene was amplified. The amplification products were digested with restriction enzymes. PCR-RFLP for MvaI restriction enzyme was observed. Frequencies of alleles were as follows: MTCYB(C)--0.926, MTCYB(G)-- 0.074. The frequencies of MTCYB/MvaI alleles found in this study for non-homing pigeons considerably deviate from the values found for homing/racing pigeons (allele MTCYB(G) occurred only in the non-homing breeds).     

Analysis of the C4 genes in baleen whales using a human cDNA probe

We have used a human C4 cDNA probe to investigate the complement component C4 gene in four members of the family Balaenopteridae: fin whale (Balaenoptera physalus), sei whale (B. borealis), minke whale (B. acutorostrata), and bryde's whale (B. edeni). Restriction mapping of genomic DNA from the first three species suggests the presence of only one locus in these species, and also shows that the C4 genes in the three species are very similar. We have used 14 restriction endonucleases to investigate the restriction fragment length polymorphism (RFLP) of fin whales, 13 enzymes for sei whales, and 8 enzymes for the minke whale. No polymorphism was seen in DNA from the five minke whale samples, but Rsa I and Taq I restriction enzymes gave polymorphism in fin and sei whales whereas Hind III and Msp I restriction enzymes showed polymorphism in sei whales only. Only one bryde's whale sample was available for investigation. The study of DNA available from mother-fetus pairs from the two polymorphic species demonstrated a simple, two-allele transmission of RFLP alleles.     

Heterogeneity in the structural basis of the human complement C4A null allele (C4A Q0) as revealed by HindIII restriction fragment length polymorphism analysis

The highly polymorphic fourth component of human complement (C4) is usually encoded by two genes. C4A and C4B, adjacent to the 21-hydroxylase (21-OH) genes, 21-OHA and 21-OHB, and is also remarkable in the high frequency of the 'null' alleles, C4A Q0 and C4B Q0. The molecular basis for the C4A Q0 allele was studied in 26 families through restriction fragment length polymorphism (RFLP) analysis with C4 and 21-OH cDNA probes after digestion of the DNA with the endonuclease HindIII. The individuals expressing the extended haplotype HLA-A1 (of A2) Cw7 B8 C2C BfS C4AQ0B1 DR3 have a large deletion taking off the C4A and 21-OHA genes.     

Lactate dehydrogenase isozymes and allozymes of the nine-spined stickleback Pungitius pungitius (L.) (Osteichthyes: Gasterosteidae)

Zymograms indicate the existence of three loci coding for lactate dehydrogenase in the nine-spined stickleback Pungitius pungitius. The skeletal muscle locus is polymorphic for two codominant alleles: Ldh-A(100). and Ldh-A(140). Differences in allele frequencies among nine samples, obtained from eight geographically isolated populations, proved not significant in 28/36 pairwise comparisons. The frequency of Ldh-A(100) varied from 0.426 to 0.715. Significant differences in apparent Km (pyruvate) values were found among the LDH-A allozymes suggesting a functional basis for the selective maintenance of this polymorphism.     

Association of AGTR1 (A1166C) and ACE (I/D) Polymorphisms with Breast Cancer Risk in North Indian Population

Renin angiotensin system (RAS) comprising Angiotensin converting enzyme (ACE), Angiotensin II (Ang II) and its receptor Angiotensin II receptor type I (AGTR1), plays a critical role in several diseases including cancer. A single nucleotide polymorphism (SNP) A1166C located in 3′ untranslated region (UTR) of AGTR1 and an insertion/deletion (I/D) polymorphism present in intron 16 of ACE gene have been associated with many diseases, but their association with Breast cancer (BCa) is still debatable. Here, we for the first time investigated the association of these polymorphisms in a North Indian BCa cohort including 161 patients and 152 healthy women. The polymorphisms were evaluated by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) respectively. The association between these polymorphisms and BCa risk was estimated by calculating Odds Ratio (OR) and chi-square (χ²) test. The DD genotype/D allele of ACE (I/D) polymorphism and “AC and CC” genotype/C allele of AGTR1 (A1166C) polymorphism were associated with higher risk of BCa when evaluated independently. Furthermore, interaction analysis of “AC and CC” and DD genotype and combination of “C and D” alleles of both polymorphisms revealed significantly greater BCa risk than that observed independently. Conclusively, women harboring “AC or CC” genotype/C allele for AGTR1 (A1166C) polymorphism and DD genotype/D allele for ACE (I/D) polymorphisms have a predisposition to develop more aggressive disease with advanced staging and larger tumor size. Our study indicates importance of genetic screening based on these polymorphisms for women, who may have higher risk of BCa.     

Detection of a new polymorphism of the human prothrombin (F2) gene by combination of PASA and mutated primer-mediated PCR-RFLP

A new polymorphism of the human prothrombin (F2) gene was detected by a combination of polymerase chain reaction (PCR) amplification of specific alleles (PASA) and mutated primer-mediated PCR restriction fragment length polymorphism (PCR-RFLP). The method is simple and useful for detecting polymorphisms and mutations. The new polymorphism of C1 and C2 examined by this method is highly heterozygous and serves as a good human DNA marker.     

Genetic polymorphism and isoenzyme patterns of lactate dehydrogenase in tench (Tinca tinca), crucian carp (Carassius carassius) and carp (Cyprinus carpio)

Isoenzyme patterns and the polymorphism of lactate dehydrogenase (LDH) were investigated in 3 fish species of family Cyprinidae, i.e. tench (Tinca tinca), crucian carp (Carassius carassius) and carp (Cyprinus carpio). The isoenzyme patterns were tissue and species specific. In crucian carp subunits with different electrophoretic mobility are present, which are genetically controlled from the B1, B2, A1, A2 and C loci, while the set of loci in carp is B1, B2, A, C1 and C2 and in tench B, A, C. The locus B of LDH in tench, the locus B2 in crucian carp, and the loci B1, C1 and C2 in carp are polymorphic and have two different alleles in each case. The polymorphism did not affect the total LDH activity in the tissues. All the populations investigated were in Hardy-Weinberg equilibrium. The genetic control of the polymorphism in B1 and C1 loci in carp was proved by test matings. The polymorphism in B loci tested in erythrocytes may be utilized as genetic markers in the fish breeding.     

中国鄂温克族和达斡尔族人群中血小板膜糖蛋白Ib(GPIb)的多态性

研究旨在进一步探索同一人种的不同民族之间GPIb的多态性是否存在差异。血小板从84名健康鄂温克族,85名达斡尔族青年志愿采血分离而获得,血小板样品经SDS-聚丙烯酰胺凝胶电泳（SDS-PAGE）后,再转移至硝酸纤维素膜上（Western blotting),用生物素化的素胚凝集素（WGA/Bio)偶联辣根过氧化酶复合物试剂盒（ABCkit)分步孵育,再以4-氯-1-萘酚显影,膜上呈现的深灰色带即是GPIbα链,其分子量分别为141、136、132、128kD,此即A、B、C、D四型,此为基因型,其中每两型构成一个表型,本研究证明,在我国鄂温克族人群中,等位基因A、B、C、D的频率分别为0.113,0.375,0.381,0.131;在达斡尔族中,A型为0.124,B型为0.406,C型为0.359。D型为0.112。经统计学分析。在这两个民族的人群之间,其GPIb各基因频率无显性差异。两个民族人群的GPIb表型经卡方检验也无显性差异。由此结果说明,鄂温克,达斡尔两个民族的人群之间,GPIb的多态性特征是相似的。     

The Ca2+-sensing receptor gene (PCAR1) mutation T151M in isolated autosomal dominant hypoparathyroidism

Isolated autosomal dominant hypoparathyroidism is a heterogeneous disorder characterized by parathyroid hormone (PTH) deficiency, hypocalcemia and hyperphosphatemia. The candidate gene approach was used to study a large Norwegian family. The loci for the PTH gene, PTH receptor gene and RET protooncogene were excluded using dinucleotide markers and restriction fragment length polymorphism analysis. Complete cosegregation of this trait was found with the chromosomal region 3q13, using the short tandem repeat markers D3S1267, D3S1269, D3S1303, D3S1518, and RHO. This region contains the candidate locus for the Ca²⁺-sensing receptor (PCAR1). By single-strand conformation polymorphism (SSCP) analysis of all PCAR1 exons followed by automated sequencing, we identified a C to T transition in exon 2 (cDNA position 452) on the mutant allele in the family. The mutation predicts a substitution of Thr to Met in amino acid position 151 (T151M). A StyI restriction site created by the nucleotide substitution was used to confirm the mutation on all alleles, as well as to exclude it among 100 normal alleles (blood donors). SSCP analysis also identified a novel polymorphism of PCAR1 intron 4 (1609–88t→c) on normal alleles.The T151M mutation is located in the extracellular N-terminal domain of PCAR1, which belongs to the superfamily of G protein-coupled receptors. We suggest that this is a gain-of-function mutation that increases the sensitivity of the receptor to [Ca²⁺], thereby decreasing the calcium set point. Received: 29 September 1995 / Revised: 19 January 1996     

Genetic variation of the sixth component of complement (C6) in the Manx shearwater (Puffinus p. puffinus)

A polymorphism in the sixth complement component (C6) has been investigated by isoelectric focusing. The results indicate that C6 is coded by genes at a single autosomal locus. Six structural alleles have been identified, a null allele is also postulated. Five island colonies of Shearwater were investigated, two alleles C6 A and C6 B predominate in all sites. The ability of rabbit C5 and C7 to combine successfully with avian C6 indicates the functional sites of these molecules are highly conserved.     

Genetic variation of the sixth component of complement (C6) in the Manx shearwater (Puffinus p. puffinus)

A polymorphism in the sixth complement component (C6) has been investigated by isoelectric focusing. The results indicate that C6 is coded by genes at a single autosomal locus. Six structural alleles have been identified, a null allele is also postulated. Five island colonies of Shearwater were investigated, two alleles C6 A and C6 B predominate in all sites.
The ability of rabbit C5 and C7 to combine successfully with avian C6 indicates the functional sites of these molecules are highly conserved.     

Identification of Novel Allelic Variants of Integrin Beta 2 (ITGB2) Gene and Screening for Bubaline Leukocyte Adhesion Deficiency Syndrome in Indian Water Buffaloes (Bubalus bubalis)

A fragment of 570 bp corresponding to exon 5 and 6 of integrin beta 2 (ITGB2) gene was amplified for screening D128G mutation in one hundred and fifty two buffaloes (Bubalus bubalis) which causes bovine leukocyte adhesion deficiency syndrome (BLAD) in cattle, as well as to ascertain polymorphism. TaqI PCR-RFLP revealed no such mutation thus indicating the absence of bubaline leukocyte adhesion deficiency (BuLAD) allele in animals under study. However, the polymorphism studies using MspI restriction enzyme revealed two genotypic patterns viz. AA pattern (bands of 293, 141, 105, and 31 bp) and BB pattern (bands of 293, 105, 77, 64, and 31 bp). The sequences of A and B alleles were submitted to the GenBank (EU853307 and AY821799).     

Restriction fragment length polymorphism (RFLP) of exon 2 of the MhcBibi-DRB3 gene in American bison (Bison bison)

Polymerase chain reaction (PCR) primers specific to exon 2 of the bovine lymphocyte antigen (BoLA)-DRB3 gene were successfully used to amplify the equivalent region in 469 American bison (Bison bison). In domestic cattle, alleles of DRB3 are assigned through a restriction fragment length polymorphism (RFLP) analysis of the patterns of fragment lengths observed after digestion with the restriction enzymes RsaI, BstYI and HaeIII. In bison, using the same procedure, the observed RFLP patterns provided evidence for the strong conservation of restriction sites previously reported in cattle.     

Molecular characterization of genetic mutation in human lactate dehydrogenase-A (M) deficiency

Human lactate dehydrogenase-A mutant gene was isolated from the genomic DNA library of a patient deficient in LDH-A (Muscle) subunit. The nucleotide sequences of seven protein-coding exons were determined and a deletion of 20 base-pairs in exon 6 was found. This mutation results in a frame-shift translation and premature termination. The predicted incomplete LDH-A (M) subunit containing only 259 instead of 331 amino acids appears to be degraded rapidly, since no protein was detected immunologically (Maekawa et al., Am J Hum Genet 39:232-238, 1986). In addition, three synonymous (silent) substitutions, A to C, T to C, and G to A, were observed at codons 115, 160 and 172, respectively, in this LDH-A mutant gene.     

Analysis of genetic mutations in human lactate dehydrogenase-A(M) deficiency using DNA conformation polymorphism in combination with polyacrylamide gradient gel and silver staining

Human lactate dehydrogenase (LDH)-A mutant gene was analyzed by polymerase chain reaction - DNA conformation polymorphism (DCP). We used polyacrylamide gradient gel and silver staining procedures for DCP analysis and observed abnormal migration patterns in individuals heterozygous for LDH-A deficiency. Further sequence determination of the mutant alleles consistently resulted in detection of base substitutions, a G to T transversion at codon 328 (GAG----TAG), and synonymous substitutions at codon 115, 160 and 172. Such mutations were easily detectable using the DCP technique. The DCP technique using the polyacrylamide gradient gel and silver staining method seems likely to be useful for the rapid screening of mutations and for further genotype detection.     

Affinity differences for vitamin D metabolites associated with the genetic isoforms of the human serum carrier protein (DBP)

Human vitamin D binding protein (DBP) displays considerable polymorphism with 120 described alleles. Among these, three alleles are frequently observed, Gc 1F (pI 4.94–4.84), Gc 1S (pI 4.95–4.85) and Gc 2 (pI 5.1). Differences between these genetic forms of the protein in affinity for vitamin D metabolites have been detected by electrophoretic methods. The constant affinity (Ka) values determined in this study confirm these differences. The affinities of six rare variants were also examine. Those of the DBP genetic forms to the vitamin D derivatives 25-OH-D₃ and 1,25-(OH)₂-D₃ seem to be related to the isoelectric point of the proteins: a high affinity corresponding to a low isoelectric point. The Gc 1A9 and 1A11 mutants were associated with higher affinity for the vitamin D derivatives and the Gc 1C1 and 1C21 mutants were deficient.     

Association analysis of gene polymorphism in alcohol metabolizing enzymes with risk for coronary atherosclerosis

The allele and genotype distribution of two alcohol dehydrogenase genes ADH1B (exon 3 polymorphism A/G (47His)), ADH7 (intron 5 polymorphism G/C) and cytochrome P450 2E1 gene (CYP2E1; 5'-flanking region G/C and intron 6 T/A polymorphisms) were examined in Russian (Tomsk, n = 125) healthy population and in coronary atherosclerosis patients (CA, n = 92). The genotype frequencies followed the Hardy-Weinberg equilibrium and the alleles were in linkage equilibrium or gametic equilibrium in the control sample. Only two CYP2E1 gene polymorphisms were in linkage disequilibrium. The frequencies of the derived alleles at ADH1B (*G (+MslI) allele), CYP2E1 (**C2 (+PstI) allele) and CYP2E1 (*C (-Dra I)2 allele) were 8.48 +/- 1.86%; 1.20 +/- 0.69% and 10.00 +/- 1.90%, respectively. The 2ADH7 gene polymorphism showed a high level of heterozygosity; the frequency of the ADH7*C (-Sty I) allele was 44.58 +/- 3.21%. A significantly higher frequency of CYP2E1 (*C2 (+Pst I)) allele has been revealed in the CA group (P = 0.043; OR = 4.23; 95% CI 1.03-20.01). The tendency to significant effect of A1A2 genotype in ADH1B Msl 1 polymorphism was observed for systolic blood pressure in the control group (P = 0.068). The statistically significant two-way interaction effects of ADH7 StyI and CYP2E1 DraI on diastolic blood pressure (P = 0.029) and on the serum high density lipoprotein level (P = 0,042) were also revealed. Association of A1A2 genotype in ADHIB Msl I polymorphism with reduced amount in a serum of a very low density lipoprotein level (P = 0.045) have also been shown. This may result from multifunctional activity of alcohol metabolizing enzymes and their involvement in many metabolic and free radical reactions in the body.