ABSTRACT
The most common mutation in alpha-1-antitrypsin deficiency, conversion of a G to an A at base 9989 (PI-Z), was detected with the chemical cleavage of mismatch method, demonstrating the power of the method for prenatal diagnosis. Exon V of the gene was amplified using the polymerase chain reaction and heteroduplexes were formed to test for the presence of the mutation. The predicted C mismatch was readily detectable with hydroxylamine, and by making the probe from the chorionic villus sample it was possible to determine that the fetus was heterozygous, not homozygous, for the mutation.
Subject(s)
Prenatal Diagnosis/methods , alpha 1-Antitrypsin Deficiency , Autoradiography , Base Sequence , Chorionic Villi Sampling , Female , Humans , Hydroxylamine , Hydroxylamines , Molecular Sequence Data , Mutation , Polymerase Chain Reaction , Pregnancy , alpha 1-Antitrypsin/geneticsABSTRACT
A single base substitution is responsible for the PI-Z mutation in alpha-1-antitrypsin (AAT) deficiency. The Z mutation, which is in exon V of the AAT gene, was analysed directly using a primer designed with a single base substitution in the DNA sequence. During the polymerase chain reaction with this primer, a restriction enzyme site was created in the exon-V-amplified DNA sequence; this site was present in the normal allele (M form) but absent in the Z form. Here, the design of the primer and the application of the designer primer for prenatal diagnosis of chorion villus samples (CVS) for AAT deficiency is described. The method provides a simple rapid means of prenatal diagnosis of AAT deficiency within a day of the collection of the CVS. The detection of the nucleotide base change in AAT deficiency at the Z mutation site provides the opportunity for accurate prenatal diagnosis where no tissue is available from an AAT-affected individual.
Subject(s)
Polymerase Chain Reaction , alpha 1-Antitrypsin Deficiency , Base Sequence , DNA , Deoxyribonucleases, Type II Site-Specific/metabolism , Female , Humans , Male , Molecular Sequence Data , Pedigree , alpha 1-Antitrypsin/geneticsABSTRACT
Cystic fibrosis is a common autosomal recessive disease in white persons. Prenatal diagnosis by DNA analysis became possible in families with a child who is affected by cystic fibrosis when the probes pJ3.11, metH and metD, which are linked closely to the cystic fibrosis gene (CF) were described. The recent description of the XV-2c and KM.19 probes has improved the prenatal diagnosis of cystic fibrosis greatly. The KM.19 probe alone was informative in eight of 12 families that were studied while XV-2c was informative in eight of 12 families that were studied while XV-2c was informative in only two of the 12 families. In contrast, the use of the pJ3.11, metH and metD probes in combination allowed full diagnosis in six of the 12 families. The combined use of the CF-linked probes produced informative data for all 12 families. Therefore, in most families with at least one affected living child, the first-trimester diagnosis of cystic fibrosis is possible with fetal DNA that has been prepared from chorionic villous samples. Strong linkage disequilibrium was found with both the KM.19-PstI polymorphism and the XV-2c-TaqI polymorphism and the CF gene.
Subject(s)
Cystic Fibrosis/diagnosis , DNA Probes/analysis , Fetal Diseases/diagnosis , Genetic Linkage , Prenatal Diagnosis , Alleles , Cystic Fibrosis/epidemiology , Cystic Fibrosis/genetics , DNA Probes/classification , Female , Fetal Diseases/epidemiology , Fetal Diseases/genetics , Genetic Counseling , Genotype , Haplotypes , Humans , Male , Polymorphism, Restriction Fragment Length , Pregnancy , Pregnancy Trimester, First , Probability , Recombination, Genetic , VictoriaSubject(s)
Disease Susceptibility/immunology , Histocompatibility Antigens Class II/immunology , Major Histocompatibility Complex , Antibodies, Monoclonal , Autoimmune Diseases/immunology , Basement Membrane/physiology , Cells, Cultured , Endothelium/immunology , Humans , Immunity, Cellular , Immunization , Lymphocytes/immunology , Monocytes/immunology , Racial GroupsABSTRACT
The pathogenesis of insulin-dependent diabetes mellitus (IDDM) will remain obscure until the number of genetic mechanisms contributing to susceptibility can be clarified. Australian multiple-case families of IDDM have been examined for concordance in IDDM and HLA haplotypes and analysed for goodness-of-fit to hypotheses of one or two high-risk susceptibility genes. Diabetic siblings are HLA-identical in 75% of cases, confirming the association between HLA and IDDM, and suggesting recessively in inheritance of IDDM susceptibility. However, the most striking finding is that 52% of IDDM offspring are positive for both HLA-DRW3 and DRW4, compared with only 8% of their non-diabetic sibs and 1% of the general population. The risk for IDDM for the HLA-DRW3/DRW4 heterozygote is 37.2, and the chance that a child from a multiple-case family of IDDM will himself develop the disease is 6.5 times as great if he is a HLA-DRW3/DRW4 heterozygote than if he is not positive for both antigens. Possible genetic mechanisms are discussed, but the present data strongly support the interaction of two HLA-DR associated susceptibility genes in IDDM and rejects the hypothesis of a single autosomal recessive susceptibility gene.
