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Molecular genetic studies

Molecular genetic studies

The purpose of carrying out molecular genetic methods of investigation is to determine the presence of modifications and changes in some chromosomes, DNA sections or genes.Widely used in practice, this method of working with DNA was received in the 70-80's.The last century.

Table of contents:
  1. MonogenicGenetic diseases;
  2. Probability of development of oncological diseases;
  3. Presence of factors provoking multifactorial diseases.

Determining the risk of development of onco processes using molecular genetic research reveals :

  • Risk of developing gastric and thyroid cancer;
  • Probability of colon cancer and early stages of the disease;
  • Genetic predisposition to the development of cancer of the body of the uterus, ovaries, milk and prostate;
  • Presence of recombination of ABL / BCR genes detected in leukemia;
  • Presence of prerequisites ensuring the effectiveness of antitumor therapy with hefatinib in the presence of non-small cell cancer.

Conducting molecular genetic tests for the presence of genetically determined prerequisites for the development of multifactorial diseases, it is possible to identify the risk of development:

  • hypertension;
  • pre-eclampsia;
  • rheumatoid arthritis;
  • of osteoporosis;
  • violations of lipid metabolism;
  • of type 1 and type 2 diabetes;
  • diseases of the reproductive system.

With the help of this method, the metabolism and the justification of the use of certain medications are evaluated.

To whom are assigned molecular genetic studies

Molecular genetic tests are shown to persons:

  • suffering from infertility;
  • exposed to adverse environmental factors;
  • has close relatives in the family, suffering from oncological, mental, vascular and endocrine diseases.

How is the molecular genetic test performed?

How is the molecular genetic test performed

The first stage of molecular genetic analysis is very important and consists in obtaining RNA and DNA samples that are individual fragments of the cell's DNA or its entire chain.To select the required number of fragments, a method of amplification, i.e., multiplying them by polymerase chain reaction( enzymatic replication) is used.

For the analysis of DNA molecules, their preliminary division into parts and treatment with bacterial endonucleases( restriction enzymes) - enzymes, which can cut a double helix of DNA into parts of 4-6 pairs, is required.

DNA fragments are separated by length and size with a special gel( polyacrylamide and agarose), using electrophoresis.Under the action of the latter, they move down the gel at different speeds, leaving behind a discrete strip.

Molecular genetic studies of hereditary pathologies are also used to study the human genome.The Southern blot hybridization allows, in this case, to determine the specific DNA fragments required for this.In this case, DNA denaturation is first resorted, as a result of which the fragments are obtained as a single chain and transferred to a filter( nylon or nitrocellulose), which is soaked in a buffer solution.

The gel containing the DNA fragments is transferred to a filter paper with a saline solution( with a high% concentration).A nitrocellulose filter and filter paper, but dry paper, are superimposed on top( to absorb saline solution).As a result, single-stranded DNA remains on the filter in the same position as on the gel.

To identify the necessary fragments, DNA hybridization is carried out with a cloned fragment or a radioactive DNA probe.The result of this procedure is detected by radioautography, thanks to which all the complementary probes of the DNA sequence are reflected in the form of a radioactive band.

Image 3258 The Southern method allows you to recreate the restriction map of the human genome in a specific part of the gene.This makes it possible to detect the presence of any defects in the gene itself.The developed methods are considered to be quite effective and allow for the accurate diagnosis of hereditary diseases.To this end, from the embryonic cells that are contained in the amniotic fluid, DNA is isolated.Subsequently, it is hybridized using Southern blotting with a radioactive DNA probe.As a result, it is very easy to recognize an abnormal embryo, because its DNA hybridizes exclusively with the DNA probe, which is complementary to the mutant sequence.

Modern science uses a number of methods to identify mutations.All of them are divided into indirect and direct molecular genetic methods of research.

Indirect methods for detecting mutations are used if the position of the gene on the genetic map is known, but its nucleotide sequence is not deciphered.

Direct diagnostics can occur in several forms:

  1. Sequencing.This is a technique for detecting the nucleotide sequence for determining the substitution of bases in a specific fragment.
  2. Blot hybridization but to Southern.This is a restrictive analysis, with the help of which mutations having a violation of the restriction site are found.
  3. Allele-specific hybridization with synthetic probe.This method also makes it possible to identify mutations in genomic DNA.
  4. Electrophoresis of double-stranded DNA in a gel( uniformly denaturing, neutral).It is the splitting of DNA at the chemical and enzymatic level.In those places where the bases are incorrectly sewn, the mutation group is usually determined.
  5. Study of the electrophoretic mobility of DNA-mutant molecules.
  6. Analysis of the synthesized protein by electrophoresis.The presence of mutations is judged by the change in protein mobility in the in vitro system.

Also, mutations are diagnosed by determining polymorphic fragments( restriction lengths) in the genome.For this, the same blot hybridization technique is used in the Southern.

Among other types of polymorphism of DNA, microsatellites are also isolated.They are short DNA sequences( tandemly repeated mono-, di-, tri-, and tetranucleotide).They serve as markers of defective mutations or marker loci of allelic variants of the gene in the study.

The gene responsible for the development of Huntington's chorea, a serious pathology, was discovered in 1993. With this disease, there is a decrease in intellectual development, a disorder of CNS movements in people after 40 years.The disease is hereditary and is transmitted by an autosomal dominant type, has a 100% penetrance.The gene of the disease is located in the 4th chromosome, in the short arm.

This gene includes a nucleotide sequence in the form of repeated repetition of the nucleotide of the CAG.In healthy people, such repetitions are normal at 11-34, patients with chorea have 37-86, but usually 45. This implies that Huntington's chorea is a hereditary pathology with a gene mutation in a multiple increase in the number of its copies( expansion).

Viktorova Julia, obstetrician-gynecologist


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