Highly polymorphic dinucleotide repeat at the NF2 gene

A highly polymorphic CA repeat was identified in a cosmid containing the 5 end of the NF2 tumour suppressor gene. This marker has proved useful in presymptomatic diagnosis in affected families.     

The human pyruvate dehydrogenase complex: a polymorphic region of the lipoate acetyl transferase (E2) subunit gene

A major issue in the study of the pathogenesis of primary biliary cirrhosis is whether the E2 subunit of the pyruvate dehydrogenase complex (PDH-E2), the major autoantigen in the disease, exists as a tissue-specific isoform. cDNA clones spanning a segment of the 3'-catalytic region of PDH-E2 (nt 1158-1361) have been isolated from human kidney, placenta and bile epithelium cells. Nucleotide sequence analysis of the clones showed differences consistent with the presence of normal variants of PDH-E2 in the human population. However, the existence of tissue-specific isoforms of PDH-E2 cannot yet be discounted.     

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.     

Primary structure of the amino-terminal (vitamin K-dependent) region of human prothrombin

The amino acid sequence of residues 1–51 of human prothrombin fragment 1 has been determined. This 51 residue peptide is 1 residue shorter than the comparable bovine region and 8 of the amino acid residues are different. The positions of 10 glutamic acid residues are identical to the γ-carboxylated ones in the bovine species. From this homology and additional evidence, these residues in human prothrombin are considered to be γ-carboxylated. The sequence was completed by automated Edman degradation of the reduced, alkylated fragment and its subfragmentation with trypsin, thermolysin and acid hydrolysis.     

Staphylocoagulase-binding region in human prothrombin

A staphylocoagulase-binding region in human prothrombin was studied by utilizing several fragments prepared from prothrombin by limited proteolysis. Bovine prothrombin, prethrombin 1, prethrombin 2, and human diisopropylphosphorylated alpha-thrombin strongly inhibited formation of the complex ("staphylothrombin") between human prothrombin and staphylocoagulase, but bovine prothrombin fragment 1 and fragment 2 had no effect on the complex formation, indicating that the binding region of human prothrombin for staphylocoagulase is located in the prethrombin 2 molecule. To identify further the staphylocoagulase-binding region, human alpha-thrombin was cleaved into the NH2-terminal large fragment (Mr = 26,000) and the COOH-terminal fragment (Mr = 16,000) by porcine pancreatic elastase. Of these fragments, the COOH-terminal fragment, which includes Asn-200 to the COOH-terminal end of the alpha-thrombin molecule, partially inhibited the complex formation between staphylocoagulase and human prothrombin. In contrast, the NH2-terminal large fragment did not show any inhibitory effect on the staphylothrombin formation. These results suggest that the staphylocoagulase interacts with human prothrombin through the COOH-terminal region of alpha-thrombin B chain. Other plasma proteins, factor X, factor IX, protein C, protein S, protein Z, all of which are structurally similar to prothrombin, did not inhibit the staphylothrombin formation at all, indicating that a specific interaction site with staphylocoagulase is contained only in the prothrombin molecule.     

Analysis of a 1963-bp polymorphic region flanking the human insulin gene

The nucleotide sequence of a long polymorphic region located 365 bp upstream from the human insulin gene is reported. The region is composed of 139 repeating sequences whose consensus structure is related to ACAGGGGTGTGGGG. Expansion in the number of repeating sequences appears to have taken place through duplication and triplication of 112–141-bp regions. However, ancestral polymorphic regions containing additions or deletions of 50 bp or more were not detected in two previous generations.     

Highly polymorphic DNA sequences in the distal region of the long arm of human chromosome 18

Highly polymorphic repeated DNA sequences were detected at the end of the long arm of human chromosome 18 by a recombinant DNA probe containing 56 bp of human DNA (pERT25). This was shown by hybridization of pERT25 to DNA from a panel of human X rodent somatic cell hybrids, by dot blot hybridization to flow-sorted human chromosomes, and by in situ hybridization to metaphase chromosomes. The high degree of polymorphism detected by this 18q DNA fragment makes it potentially useful for various applications, including investigations into the genetics of trisomy 18 (Edwards syndrome), linkage studies, and paternity testing.     

SstII polymorphic sites in the promoter region of the human cystatin C gene

Summary By direct sequencing of polymerase chain reaction (PCR) amplified DNA from different individuals, three point mutations have been found in a 220-bp fragment from the promoter region of the human cystatin C gene. The three mutations are all localized within a short segment of 85 bp on the same allele. One of the base substitutions results in the generation of a novel SstII restriction site and another in the loss of the commonly occurring SstII restriction site. A PCR-based assay for analysis of the two SstII sites was designed and used to demonstrate Mendelian inheritance of the polymorphism. This SstII restriction fragment lenght polymorphism offers a new probe-independent marker for chromosome 20 linkage studies.     

Nucleotide sequence of the gene for human prothrombin

A human genomic DNA library was screened for the gene coding for human prothrombin with a cDNA coding for the human protein. Eighty-one positive lambda phage were identified, and three were chosen for further characterization. These three phage hybridized with 5' and/or 3' probes prepared from the prothrombin cDNA. The complete DNA sequence of 21 kilobases of the human prothrombin gene was determined and included a 4.9-kilobase region that was previously sequenced. The gene for human prothrombin contains 14 exons separated by 13 intervening sequences. The exons range in size from 25 to 315 base pairs, while the introns range from 84 to 9447 base pairs. Ninety percent of the gene is composed of intervening sequence. All the intron splice junctions are consistent with sequences found in other eukaryotic genes, except for the presence of GC rather than GT on the 5' end of intervening sequence L. Thirty copies of Alu repetitive DNA and two copies of partial KpnI repeats were identified in clusters within several of the intervening sequences, and these repeats represent 40% of the DNA sequence of the gene. The size, distribution, and sequence homology of the introns within the gene were then compared to those of the genes for the other vitamin K dependent proteins and several other serine proteases.     

A highly polymorphic region 3'' to the human type II collagen gene.

We have characterised a highly polymorphic region 1.3kb downstream of the human Type II collagen gene. It consists of a highly AT-rich tandem repetitive region (minisatellite) approximately 650bp long. Two alleles had been observed previously, differing in size by approximately 300bp. When this region was cloned from four unrelated individuals carrying the larger allele, DNA sequence data identified three alleles, suggesting far higher polymorphism than was originally supposed. This minisatellite was shown to be present in a single copy in the human genome, and to have arisen after the divergence of Old and New World monkeys.