Many congenital metabolic disorders are caused by various enzyme defects resulting from mutations that change their structure. The use of modern biochemical methods allow to determine any metabolites specific to a specific hereditary disease. The subject of modern biochemical diagnostics are specific metabolites, enzymopathy, various proteins. Objects of biochemical analysis can be urine, sweat, plasma and serum. For biochemical diagnostics, both simple qualitative reactions and more precise methods are used. For example, a thin layer chromatography of urine and blood can be used to diagnose metabolic disorders of amino acids, oligosaccharides, mucopolysaccharides. Gas chromatography is used to detect metabolic disorders of organic acids.
Biochemical methods are also used to diagnose heterozygous conditions in adults. It is known that among healthy people there is always a large number of carriers of the pathological gene. Although such people are outwardly healthy, the probability of disease in their child always exists. In this regard, the detection of heterozygous conditions is an important task of medical genetics. If heterozygous carriers of any disease marry, the risk of giving birth to a sick child in such a family will be 25%. The chances of meeting two carriers of the same pathological gene are higher if relatives get married, ie they can inherit the same recessive gene from their common ancestor. Detection of heterozygous carriers of this or that disease is possible through the use of biochemical tests, microscopic examination of blood cells and tissues, determination; activity of the enzyme changed as a result of mutation. It is known that diseases based on metabolic disorders form a significant part of hereditary pathology. Thus, heterozygous carriers of phenylketonurine react to the introduction of phenylalanine with a higher amino acid content in plasma than normal homozygotes.
The biochemical method is widely used in medical genetic counseling to determine the risk of giving birth to a sick child. Advances in biochemical genetics contribute to the wider introduction of diagnosis of heterozygous carriers in practice. Until recently, it was possible to diagnose no more than 10-15 heterozygous conditions, at present – more than 200. However, it should be noted that there are still many hereditary diseases for which methods of heterozygous diagnostics have not yet been developed.
Molecular and genetic method
Molecular genetic method – detection of changes in certain areas of DNA, gene or chromosome. It is based on modern methods of working with DNA or RNA, in 70-80 due to progress in molecular genetics and success in studying the human genome molecular genetic approach has found wide application, The initial stage of molecular genetic analysis is to obtain DNA or RNA samples. For this purpose, genomic DNA (all DNA of a cell) or its individual fragments are used. In the latter case, in order to obtain a sufficient number of such fragments, it is necessary to amplify (multiply) them, using polymerase chain reaction – a fast method of enzymatic replication of a certain DNA fragment. It can be used to amplify any DNA fragment located between two known sequences. It is impossible to analyze huge DNA molecules as they exist in a cell. These enzymes are capable of cutting a double helix of DNA, and the cutting locations are strictly specific to the sample. The DNA cleavage by restrictases provides a characteristic set of fragments (4-6 base pairs) that differ in length. Fractionation of DNA fragments by size and length is carried out with electrophoresis on the surface of agarose or acrylamide gel. Under the action of an electric field, the DNA fragments begin to move down the gel at a rate that depends on their length. As a result, each DNA fragment occupies a certain position in the form of a discrete band at a specific location of the gel. The length of each fragment can be determined by comparing the distance traveled by the fragment with a standard DNA segment.
Molecular genetic diagnosis of hereditary diseases is also used to study the human genome, and blot hybridization by Sauserne is used to identify the necessary specific DNA fragments. The essence of this technique is as follows: first, DNA denaturation is performed to form single-chain fragments that are transferred to a nitrocellulose or nylon filter in a buffer solution.