What does molecular biology study in brief? Molecular biology. Molecular Biology Methods

1. Introduction.

Subject, tasks and methods of molecular biology and genetics. The importance of “classical” genetics and genetics of microorganisms in the development of molecular biology and genetic engineering. The concept of a gene in “classical” and molecular genetics, its evolution. Contribution of genetic engineering methodology to the development of molecular genetics. Applied significance of genetic engineering for biotechnology.

2. Molecular basis of heredity.

The concept of a cell, its macromolecular composition. The nature of the genetic material. The history of evidence for the genetic function of DNA.

2.1. Various types of nucleic acids. Biological functions of nucleic acids. Chemical structure, spatial structure and physical properties nucleic acids. Features of the structure of genetic material of pro- and eukaryotes. Complementary Watson-Crick base pairs. Genetic code. The history of deciphering the genetic code. Basic properties of the code: tripletity, code without commas, degeneracy. Features of the code dictionary, codon families, semantic and “nonsense” codons. Circular DNA molecules and the concept of DNA supercoiling. DNA topoisomers and their types. Mechanisms of action of topoisomerases. Bacterial DNA gyrase.

2.2. DNA transcription. Prokaryotic RNA polymerase, its subunit and three-dimensional structures. Variety of sigma factors. Prokaryotic gene promoter, its structural elements. Stages of the transcription cycle. Initiation, formation of an “open complex,” elongation and termination of transcription. Transcription attenuation. Regulation of tryptophan operon expression. “Riboswitches.” Mechanisms of transcription termination. Negative and positive regulation of transcription. Lactose operon. Transcription regulation in lambda phage development. Principles of DNA recognition by regulatory proteins (CAP protein and lambda phage repressor). Features of transcription in eukaryotes. RNA processing in eukaryotes. Capping, splicing and polyadenylation of transcripts. Splicing mechanisms. The role of small nuclear RNAs and protein factors. Alternative splicing, examples.

2.3. Broadcast, its stages, ribosome function. Localization of ribosomes in the cell. Prokaryotic and eukaryotic types of ribosomes; 70S and 80S ribosomes. Morphology of ribosomes. Division into subparticles (subunits). Codon-dependent aminoacyl-tRNA binding in the elongation cycle. Codon-anticodon interaction. Involvement of elongation factor EF1 (EF-Tu) in the binding of aminoacyl-tRNA to the ribosome. Elongation factor EF1B (EF-Ts), its function, sequence of reactions with its participation. Antibiotics that act on the stage of codon-dependent binding of aminoacyl-tRNA to the ribosome. Aminoglycoside antibiotics (streptomycin, neomycin, kanamycin, gentamicin, etc.), their mechanism of action. Tetracyclines as inhibitors of aminoacyl-tRNA binding to the ribosome. Initiation of broadcast. Main stages of the initiation process. Translation initiation in prokaryotes: initiation factors, initiation codons, 3¢ end of small ribosomal subunit RNA and Shine-Dalgarno sequence in mRNA. Translation initiation in eukaryotes: initiation factors, initiation codons, 5¢ untranslated region and cap-dependent “terminal” initiation. “Internal” cap-independent initiation in eukaryotes. Transpeptidation. Transpeptidation inhibitors: chloramphenicol, lincomycin, amycetin, streptogramins, anisomycin. Translocation. Involvement of elongation factor EF2 (EF-G) and GTP. Translocation inhibitors: fusidic acid, viomycin, their mechanisms of action. Termination of broadcast. Stop codons. Protein termination factors of prokaryotes and eukaryotes; two classes of termination factors and their mechanisms of action. Regulation of translation in prokaryotes.

2.4. DNA replication and its genetic control. Polymerases involved in replication, characteristics of their enzymatic activities. Accuracy of DNA reproduction. The role of steric interactions between DNA base pairs during replication. E. coli polymerases I, II and III. Polymerase III subunits. Replication fork, “leading” and “lagging” strands during replication. Fragments of Okazaki. A complex of proteins at the replication fork. Regulation of replication initiation in E. coli. Termination of replication in bacteria. Features of the regulation of plasmid replication. Bidirectional and rolling circle replication.

2.5. Recombination, its types and models. General or homologous recombination. DNA double-strand breaks that initiate recombination. The role of recombination in post-replicative repair of double-strand breaks. Holliday structure in the recombination model. Enzymology of general recombination in E. coli. RecBCD complex. RecA protein. The role of recombination in ensuring DNA synthesis during DNA damage that interrupts replication. Recombination in eukaryotes. Recombination enzymes in eukaryotes. Site-specific recombination. Differences in the molecular mechanisms of general and site-specific recombination. Classification of recombinases. Types of chromosomal rearrangements carried out during site-specific recombination. Regulatory role of site-specific recombination in bacteria. Construction of chromosomes of multicellular eukaryotes using a phage site-specific recombination system.

2.6. DNA repair. Classification of types of reparation. Direct repair of thymine dimers and methylated guanine. Cutting out the bases. Glycosylases. The mechanism of repair of unpaired nucleotides (mismatch repair). Selecting the DNA strand to be repaired. SOS reparation. Properties of DNA polymerases involved in SOS repair in prokaryotes and eukaryotes. The concept of “adaptive mutations” in bacteria. Repair of double-strand breaks: homologous post-replicative recombination and joining of non-homologous ends of the DNA molecule. The relationship between the processes of replication, recombination and repair.

3. Mutation process.

The role of biochemical mutants in the formation of the one gene – one enzyme theory. Classification of mutations. Point mutations and chromosomal rearrangements, the mechanism of their formation. Spontaneous and induced mutagenesis. Classification of mutagens. Molecular mechanism of mutagenesis. The relationship between mutagenesis and repair. Identification and selection of mutants. Suppression: intragenic, intergenic and phenotypic.

4. Extrachromosomal genetic elements.

Plasmids, their structure and classification. Sex factor F, its structure and life cycle. The role of factor F in the mobilization of chromosomal transfer. Formation of donors of the Hfr and F types." The mechanism of conjugation. Bacteriophages, their structure and life cycle. Virulent and temperate bacteriophages. Lysogeny and transduction. General and specific transduction. Migrating genetic elements: transposons and IS sequences, their role in genetic exchange. DNA -transposons in the genomes of prokaryotes and eukaryotes. IS sequences of bacteria, their structure. IS sequences as a component of the F-factor of bacteria, which determines the ability of transfer of genetic material during conjugation. Direct non-replicative and replicative mechanisms of transposition. transfer of transposons and their role in structural rearrangements (ectopic recombination) and in genome evolution.

5. Study of gene structure and function.

Elements of genetic analysis. Cis-trans complementation test. Genetic mapping using conjugation, transduction and transformation. Construction of genetic maps. Fine genetic mapping. Physical analysis of gene structure. Heteroduplex analysis. Restriction analysis. Sequencing methods. Polymerase chain reaction. Identification of gene function.

6. Regulation of gene expression. Concepts of operon and regulon. Control at the level of transcription initiation. Promoter, operator and regulatory proteins. Positive and negative control of gene expression. Control at the level of transcription termination. Catabolite-controlled operons: models of lactose, galactose, arabinose and maltose operons. Attenuator-controlled operons: a model of the tryptophan operon. Multivalent regulation of gene expression. Global regulatory systems. Regulatory response to stress. Posttranscriptional control. Signal transduction. Regulation involving RNA: small RNAs, sensor RNAs.

7. Basics of genetic engineering. Restriction and modification enzymes. Isolation and cloning of genes. Vectors for molecular cloning. Principles of designing recombinant DNA and their introduction into recipient cells. Applied aspects of genetic engineering.

A). Main literature:

1. Watson J., Tooze J., Recombinant DNA: A Short Course. – M.: Mir, 1986.

2. Genes. – M.: Mir. 1987.

3. Molecular biology: structure and biosynthesis of nucleic acids. / Ed. . – M. Higher school. 1990.

4. – Molecular biotechnology. M. 2002.

5. Spirin ribosomes and protein biosynthesis. – M.: Higher School, 1986.

b). Additional literature:

1. Hesin genome. – M.: Science. 1984.

2. Rybchin genetic engineering. – St. Petersburg: St. Petersburg State Technical University. 1999.

3. Patrushev genes. – M.: Nauka, 2000.

4. Modern microbiology. Prokaryotes (in 2 vols.). – M.: Mir, 2005.

5. M. Singer, P. Berg. Genes and genomes. – M.: Mir, 1998.

6. Shchelkunov engineering. – Novosibirsk: From Sib. Univ., 2004.

7. Stepanov biology. Structure and functions of proteins. – M.: V. Sh., 1996.

Molecular biology has experienced a period rapid development own research methods, which now differ from biochemistry. These include, in particular, methods of genetic engineering, cloning, artificial expression and gene knockout. Since DNA is the material carrier of genetic information, molecular biology significantly closer to genetics, and at the junction molecular genetics was formed, which is both a branch of genetics and molecular biology. Just as molecular biology widely uses viruses as a research tool, virology uses molecular biology methods to solve its problems. To analyze genetic information, we use Computer Engineering, in connection with which new areas of molecular genetics have emerged, which are sometimes considered special disciplines: bioinformatics, genomics and proteomics.

History of development

This seminal discovery was prepared by a long period of research into the genetics and biochemistry of viruses and bacteria.

In 1928, Frederick Griffith first showed that an extract of heat-killed pathogenic bacteria could transmit pathogenicity to non-dangerous bacteria. The study of bacterial transformation subsequently led to the purification of the pathogenic agent, which, contrary to expectations, turned out to be not a protein, but a nucleic acid. The nucleic acid itself is not dangerous; it only carries genes that determine the pathogenicity and other properties of the microorganism.

In the 50s of the 20th century, it was shown that bacteria have a primitive sexual process; they are capable of exchanging extrachromosomal DNA and plasmids. The discovery of plasmids, as well as transformation, formed the basis of plasmid technology, widespread in molecular biology. Another important discovery for the methodology was the discovery of bacterial viruses and bacteriophages at the beginning of the 20th century. Phages can also transfer genetic material from one bacterial cell to another. Infection of bacteria by phages leads to changes in the composition of bacterial RNA. If without phages the composition of RNA is similar to the composition of bacterial DNA, then after infection the RNA becomes more similar to the DNA of a bacteriophage. Thus, it was established that the structure of RNA is determined by the structure of DNA. In turn, the rate of protein synthesis in cells depends on the amount of RNA-protein complexes. This is how it was formulated central dogma of molecular biology: DNA ↔ RNA → protein.

The further development of molecular biology was accompanied by both the development of its methodology, in particular, the invention of a method for determining the nucleotide sequence of DNA (W. Gilbert and F. Sanger, Nobel Prize in Chemistry 1980), and new discoveries in the field of research into the structure and functioning of genes (see History of genetics). TO beginning of XXI century, data were obtained on the primary structure of all human DNA and a number of other organisms, the most important for medicine, agriculture and scientific research, which led to the emergence of several new directions in biology: genomics, bioinformatics, etc.

see also

• Molecular Biology (journal)
• Transcriptomics
• Molecular paleontology
• EMBO - European Organization of Molecular Biologists

Literature

• Singer M., Berg P. Genes and genomes. - Moscow, 1998.
• Stent G., Kalindar R. Molecular genetics. - Moscow, 1981.
• Sambrook J., Fritsch E.F., Maniatis T. Molecular Cloning. - 1989.
• Patrushev L. I. Gene expression. - M.: Nauka, 2000. - 000 p., ill. ISBN 5-02-001890-2

Links

Wikimedia Foundation.

• 2010.
• Ardatovsky district, Nizhny Novgorod region

Arzamas district of Nizhny Novgorod region

See what “Molecular biology” is in other dictionaries: MOLECULAR BIOLOGY - studies basic properties and manifestations of life at the molecular level. The most important directions in M. b. are studies of the structural and functional organization of the genetic apparatus of cells and the mechanism for the implementation of hereditary information... ...

See what “Molecular biology” is in other dictionaries:- explores the basic properties and manifestations of life at the molecular level. Finds out how and to what extent the growth and development of organisms, the storage and transmission of hereditary information, the transformation of energy in living cells and other phenomena are caused by... Big Encyclopedic Dictionary

See what “Molecular biology” is in other dictionaries: Modern encyclopedia

See what “Molecular biology” is in other dictionaries:- MOLECULAR BIOLOGY, the biological study of the structure and functioning of the MOLECULES that make up living organisms. The main areas of study include physical and Chemical properties proteins and NUCLEIC ACIDS such as DNA. see also… … Scientific and technical encyclopedic dictionary

molecular biology- a section of biology that explores the basic properties and manifestations of life at the molecular level. Finds out how and to what extent the growth and development of organisms, the storage and transmission of hereditary information, the transformation of energy in living cells and... ... Dictionary of microbiology

molecular biology- - Topics of biotechnology EN molecular biology ... Technical Translator's Guide

Molecular biology- MOLECULAR BIOLOGY, explores the basic properties and manifestations of life at the molecular level. Finds out how and to what extent the growth and development of organisms, the storage and transmission of hereditary information, the transformation of energy in living cells and... ... Illustrated Encyclopedic Dictionary

Molecular biology- a science that aims to understand the nature of life phenomena by studying biological objects and systems at a level approaching the molecular level, and in some cases reaching this limit. The ultimate goal is... ... Great Soviet Encyclopedia

See what “Molecular biology” is in other dictionaries:- studies the phenomena of life at the level of macromolecules (mainly proteins and nucleic acids) in cell-free structures (ribosomes, etc.), in viruses, as well as in cells. Purpose M. b. establishing the role and mechanism of functioning of these macromolecules based on... ... Chemical encyclopedia

molecular biology- explores the basic properties and manifestations of life at the molecular level. Finds out how and to what extent the growth and development of organisms, the storage and transmission of hereditary information, the transformation of energy in living cells and other phenomena... ... encyclopedic Dictionary

Books

• Molecular biology of cells. Collection of Problems, J. Wilson, T. Hunt. The book by American authors is an appendix to the 2nd edition of the textbook “Molecular Biology of the Cell” by B. Alberts, D. Bray, J. Lewis and others. Contains questions and tasks, the purpose of which is to deepen ...

(Molecularbiologe/-biologin)

• Type

  Profession after diploma

• Salary

  3667-5623 € per month

Molecular biologists study molecular processes as the basis of all life processes. Based on their results, they develop concepts for the use of biochemical processes, for example in medical research and diagnostics or in biotechnology. In addition, they may be involved in pharmaceutical manufacturing, product development, quality assurance or pharmaceutical consulting.

Responsibilities of a Molecular Biologist

Molecular biologists can work in different fields. For example, they concern the use of research results for production in areas such as genetic engineering, protein chemistry or pharmacology (drug discovery). In the chemical and pharmaceutical industries, they facilitate the translation of newly developed products from research into production, product marketing and user consultation.

IN scientific research molecular biologists study the chemical and physical properties of organic compounds, as well as chemical processes (in the field of cellular metabolism) in living organisms and publish the results of their research. In higher education institutions, they teach students, prepare for lectures and seminars, grade written work, and administer exams. Independent scientific activity is possible only after obtaining a master's and doctoral degrees.

Where do Molecular Biologists Work?

Molecular biologists find work, e.g.

• in research institutes, for example in the fields of science and medicine
• in higher education institutions
• in the chemical and pharmaceutical industry
• in environmental departments

Molecular Biologist Salary

The salary level that Molecular Biologists receive in Germany is

• from 3667€ to 5623€ per month

(according to various statistical offices and employment services in Germany)

Tasks and responsibilities of a Molecular Biologist in detail

What is the essence of the profession of Molecular Biologist?

Molecular biologists study molecular processes as the basis of all life processes. Based on their results, they develop concepts for the use of biochemical processes, for example in medical research and diagnostics or in biotechnology. In addition, they may be involved in pharmaceutical manufacturing, product development, quality assurance or pharmaceutical consulting.

Vocation Molecular biology

Molecular biology or molecular genetics deals with the study of the structure and biosynthesis of nucleic acids and the processes associated with the transfer and implementation of this information in the form of proteins. This makes it possible to understand painful disorders of these functions and possibly treat them using gene therapy. There are interfaces to biotechnology and genetic engineering in which simple organisms such as bacteria and yeast are engineered to make substances of pharmacological or commercial interest available in industrial scale thanks to targeted mutations.

Theory and practice of Molecular Biology

The chemical-pharmaceutical industry offers numerous areas of employment for molecular biologists. In industrial settings, they analyze biotransformation processes or develop and improve processes for the microbiological production of active ingredients and pharmaceutical intermediates. In addition, they are involved in moving newly developed products from research to production. By performing inspection tasks, they ensure that production facilities, equipment, analytical methods and all stages of production of sensitive products such as pharmaceuticals always meet the required quality standards. In addition, molecular biologists advise users on the use of new products.

Management positions often require a master's program.

Molecular Biologists in Research and Education

In the field of science and research, molecular biologists work on topics such as the recognition, transport, folding and codification of proteins in the cell. Research results that provide the basis for practical applications in various fields are published and thus made available to other scientists and students. At conferences and congresses they discuss and present the results of scientific activities. Molecular biologists conduct lectures and seminars, lead scientific work and take exams.

Independent scientific activity requires a master's and doctoral degree.

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1. Introduction. The essence of molecular biology

Studies the basics of life of organisms at the level of macromolecules. The goal of molecular biology is to establish the role and mechanisms of functioning of these macromolecules based on knowledge of their structures and properties.

Historically, molecular biology was formed during the development of areas of biochemistry that study nucleic acids and proteins. While biochemistry studies metabolism, the chemical composition of living cells, organisms and the chemical processes occurring in them, molecular biology focuses on the study of the mechanisms of transmission, reproduction and storage of genetic information.

And the object of study of molecular biology is the nucleic acids themselves - deoxyribonucleic acids (DNA), ribonucleic acids (RNA) - and proteins, as well as their macromolecular complexes - chromosomes, ribosomes, multienzyme systems that ensure the biosynthesis of proteins and nucleic acids. Molecular biology also borders on the objects of research and partially coincides with molecular genetics, virology, biochemistry and a number of other related biological sciences.

2. Historical excursion into the stages of development of molecular biology

As a separate branch of biochemistry, molecular biology began to develop in the 30s of the last century. Even then, the need arose to understand the phenomenon of life at the molecular level to study the processes of transmission and storage of genetic information. It was at that time that the task of molecular biology was established in the study of the properties, structure and interaction of proteins and nucleic acids.

The term “molecular biology” was first used in 1933 year William Astbury during the study of fibrillar proteins (collagen, blood fibrin, muscle contractile proteins). Astbury studied the relationship between molecular structure and biological, physical features these proteins. In the early days of molecular biology, RNA was considered to be a component only of plants and fungi, and DNA - only of animals. And in 1935 The discovery of pea DNA by Andrei Belozersky led to the establishment of the fact that DNA is contained in every living cell.

IN 1940 In 2009, a colossal achievement was the establishment by George Beadle and Edward Tatham of the cause-and-effect relationship between genes and proteins. The scientists’ hypothesis “One gene - one enzyme” formed the basis of the concept that the specific structure of a protein is regulated by genes. It is believed that genetic information is encoded by a special sequence of nucleotides in DNA, which regulates the primary structure of proteins. Later it was proven that many proteins have a quaternary structure. Various peptide chains take part in the formation of such structures. Based on this, the provision on the connection between the gene and the enzyme was somewhat transformed, and now it sounds like “One gene - one polypeptide.”

IN 1944 In 2006, American biologist Oswald Avery and his colleagues (Colin McLeod and McLean McCarthy) proved that the substance that causes the transformation of bacteria is DNA, not proteins. The experiment served as proof of the role of DNA in the transmission of hereditary information, erasing outdated knowledge about the protein nature of genes.

In the early 50s, Frederick Sanger showed that a protein chain is a unique sequence of amino acid residues. IN 1951 And 1952 years, the scientist determined the complete sequence of two polypeptide chains - bovine insulin IN(30 amino acid residues) and A(21 amino acid residues), respectively.

Around the same time, in 1951–1953 gg., Erwin Chargaff formulated rules about the ratio of nitrogenous bases in DNA. According to the rule, regardless of the species differences of living organisms in their DNA, the amount of adenine (A) is equal to the amount of thymine (T), and the amount of guanine (G) is equal to the amount of cytosine (C).

IN 1953 The genetic role of DNA has been proven. James Watson and Francis Crick, based on the X-ray diffraction pattern of DNA obtained by Rosalind Franklin and Maurice Wilkins, established the spatial structure of DNA and put forward a hypothesis, which was later confirmed, about the mechanism of its replication (duplication), which underlies heredity.

1958 year - the formation of the central dogma of molecular biology by Francis Crick: the transfer of genetic information proceeds in the direction of DNA → RNA → protein.

The essence of the dogma is that in cells there is a certain directed flow of information from DNA, which, in turn, is the original genetic text consisting of four letters: A, T, G and C. It is written in the double helix of DNA in the form sequences of these letters - nucleotides.

This text is transcribed. And the process itself is called transcription. During this process RNA is synthesized, which is identical to the genetic text, but with a difference: in RNA, instead of T, there is U (uracil).

This RNA is called messenger RNA (mRNA), or matrix (mRNA). Broadcast mRNA is carried out using the genetic code in the form of triplet sequences of nucleotides. During this process, the text of the DNA and RNA nucleic acids is converted from four-letter text to twenty-letter amino acid text.

There are only twenty natural amino acids, and there are four letters in the text of nucleic acids. Because of this, a translation from a four-letter alphabet to a twenty-letter one occurs through the genetic code, in which every three nucleotides correspond to an amino acid. So you can make as many as 64 three-letter combinations from four letters, despite the fact that there are 20 amino acids. It follows from this that the genetic code must necessarily have the property of degeneracy. However, at that time the genetic code was not known, and it had not even begun to be deciphered, but Crick had already formulated his central dogma.

Nevertheless, there was confidence that the code should exist. By that time, it had been proven that this code was triplet. This means that specifically three letters in nucleic acids ( codons) correspond to any amino acid. There are only 64 of these codons, they code for 20 amino acids. This means that each amino acid corresponds to several codons at once.

Thus, we can conclude that the central dogma is a postulate that states that a directed flow of information occurs in the cell: DNA → RNA → protein. Crick emphasized the main content of the central dogma: the reverse flow of information cannot occur, the protein is not capable of changing genetic information.

This is the main meaning of the central dogma: protein is not able to change and convert information into DNA (or RNA), the flow always goes only in one direction.

Some time after this, a new enzyme was discovered, which was not known at the time the central dogma was formulated - reverse transcriptase, which synthesizes DNA from RNA. The enzyme was discovered in viruses, which have genetic information encoded in RNA rather than DNA. Such viruses are called retroviruses. They have a viral capsule containing RNA and a special enzyme. The enzyme is a reverse transcriptase, which synthesizes DNA using the template of this viral RNA, and this DNA then serves as the genetic material for the further development of the virus in the cell.

Of course, this discovery caused great shock and much controversy among molecular biologists, since it was believed that, based on the central dogma, this could not be possible. However, Crick immediately explained that he never said it was impossible. He only said that a flow of information from protein to nucleic acids can never occur, but within nucleic acids any kind of process is quite possible: the synthesis of DNA on DNA, DNA on RNA, RNA on DNA and RNA on RNA.

Once the central dogma was formulated, a number of questions still remained: How does the four-nucleotide alphabet that makes up DNA (or RNA) code for the 20-letter alphabet of amino acids that make up proteins? What is the essence of the genetic code?

The first ideas about the existence of a genetic code were formulated by Alexander Downes ( 1952 g.) and Georgy Gamov ( 1954 G.). Scientists have shown that the nucleotide sequence must include at least three units. It was later proven that such a sequence consists of three nucleotides called codon (triplet). Nevertheless, the question of which nucleotides are responsible for the inclusion of which amino acid in a protein molecule remained open until 1961.

And in 1961 Marshall Nirenberg and Heinrich Mattei used the system to broadcast in vitro. An oligonucleotide was used as a template. It contained only uracil residues, and the peptide synthesized from it included only the amino acid phenylalanine. Thus, the meaning of the codon was established for the first time: the UUU codon encodes phenylalanine. The field of the Har Quran found that the nucleotide sequence UCUCUCUCUCUC encodes a set of amino acids serine-leucine-serine-leucine. By and large, thanks to the work of Nirenberg and Korana, to 1965 year, the genetic code was completely solved. It turned out that each triplet encodes a specific amino acid. And the order of codons determines the order of amino acids in a protein.

The main principles of the functioning of proteins and nucleic acids were formulated by the early 70s. It has been recorded that the synthesis of proteins and nucleic acids is carried out using a template mechanism. The matrix molecule carries encoded information about the sequence of amino acids or nucleotides. During replication or transcription, DNA serves as the template; during translation and reverse transcription, mRNA serves as the template.

Thus, the prerequisites were created for the formation of areas of molecular biology, including genetic engineering. And in 1972, Paul Berg and his colleagues developed molecular cloning technology. Scientists have obtained the first recombinant DNA in vitro. These outstanding discoveries formed the basis of a new direction in molecular biology, and 1972 The year has since been considered the date of birth of genetic engineering.

3. Methods of molecular biology

Enormous advances in the study of nucleic acids, the structure of DNA and protein biosynthesis have led to the creation of a number of methods that are of great importance in medicine, agriculture and science in general.

After studying the genetic code and the basic principles of storage, transmission and implementation of hereditary information, special methods became necessary for the further development of molecular biology. These methods would allow genes to be manipulated, changed and isolated.

The emergence of such methods occurred in the 1970–1980s. This gave a huge impetus to the development of molecular biology. First of all, these methods are directly related to obtaining genes and their introduction into the cells of other organisms, as well as the possibility of determining the sequence of nucleotides in genes.

3.1. DNA electrophoresis

DNA electrophoresis is a basic method of working with DNA. DNA electrophoresis is used together with almost all other methods to isolate the desired molecules and further analyze the results. The gel electrophoresis method itself is used to separate DNA fragments by length.

Before or after electrophoresis, the gel is treated with dyes that can bind to DNA. The dyes fluoresce under ultraviolet light, producing a pattern of stripes in the gel. To determine the lengths of DNA fragments, they can be compared with markers- sets of fragments of standard lengths that are applied to the same gel.

Fluorescent proteins

When studying eukaryotic organisms, it is convenient to use fluorescent proteins as marker genes. The gene for the first green fluorescent protein ( green fluorescent protein, GFP) isolated from jellyfish Aqeuorea victoria, after which they were introduced into various organisms. Afterwards, genes for fluorescent proteins of other colors were isolated: blue, yellow, red. To obtain proteins with properties of interest, such genes have been artificially modified.

In general, the most important tools for working with the DNA molecule are enzymes that carry out a number of DNA transformations in cells: DNA polymerases, DNA ligases And restriction enzymes (restriction endonucleases).

Transgenesis

Transgenesis is called the transfer of genes from one organism to another. And such organisms are called transgenic.

Recombinant protein preparations are produced by the method of gene transfer into microbial cells. Mainly such protein preparations are interferons, insulin, some protein hormones, as well as proteins for the production of a number of vaccines.

In other cases, cell cultures of eukaryotes or transgenic animals are used, mostly cattle, which secrete the necessary proteins into milk. In this way, antibodies, blood clotting factors and other proteins are obtained. The transgenesis method is used to obtain cultivated plants that are resistant to pests and herbicides, and wastewater is purified with the help of transgenic microorganisms.

In addition to all of the above, transgenic technologies are indispensable in scientific research, because the development of biology occurs faster with the use of methods of modification and gene transfer.

Restriction enzymes

The sequences recognized by restriction enzymes are symmetrical, so any kind of breaks can occur either in the middle of such a sequence or with a shift in one or both strands of the DNA molecule.

When any DNA is digested with a restriction enzyme, the sequences at the ends of the fragments will be the same. They will be able to connect again because they have complementary regions.

You can get a single molecule by stitching together these sequences using DNA ligases. Due to this, it is possible to combine fragments of two different DNAs and obtain recombinant DNA.

3.2. PCR

The method is based on the ability of DNA polymerases to complete the second strand of DNA along a complementary strand in the same way as during the process of DNA replication in a cell.

3.3. DNA sequencing

The rapid development of the sequencing method makes it possible to effectively determine the characteristics of the organism under study at the level of its genome. The main advantage of such genomic and post-genomic technologies is the increased ability to research and study the genetic nature of human diseases, in order to take the necessary measures in advance and avoid diseases.

Through large studies it is possible to obtain the necessary data on various genetic characteristics different groups people, thereby developing methods of medicine. Because of this, identifying genetic predisposition to various diseases is very popular today.

Similar methods are widely applicable almost all over the world, including in Russia. Due to scientific progress, such methods are being introduced into medical research and medical practice in general.

4. Biotechnology

Biotechnology- a discipline that studies the possibilities of using living organisms or their systems to solve technological problems, as well as creating living organisms with the desired properties through genetic engineering. Biotechnology applies methods of chemistry, microbiology, biochemistry and, of course, molecular biology.

The main directions of development of biotechnology (the principles of biotechnological processes are being introduced into the production of all industries):

1. Creation and production of new types of food and animal feed.
2. Obtaining and studying new strains of microorganisms.
3. Breeding new varieties of plants, as well as creating means to protect plants from diseases and pests.
4. Application of biotechnology methods for environmental needs. Such biotechnology methods are used for processing waste disposal, cleaning Wastewater, exhaust air and soil remediation.
5. Production of vitamins, hormones, enzymes, serums for medical needs. Biotechnologists are developing improved medications which were previously considered incurable.

A major achievement of biotechnology is genetic engineering.

Genetic Engineering- a set of technologies and methods for obtaining recombinant RNA and DNA molecules, isolating individual genes from cells, manipulating genes and introducing them into other organisms (bacteria, yeast, mammals). Such organisms are capable of producing final products with the desired, modified properties.

Genetic engineering methods are aimed at constructing new, previously non-existent combinations of genes in nature.

Speaking about the achievements of genetic engineering, it is impossible not to touch on the topic of cloning. Cloning is a method of biotechnology used to produce identical offspring of different organisms through asexual reproduction.

In other words, cloning can be thought of as the process of creating genetically identical copies of an organism or cell. And cloned organisms are similar or even identical not only in external signs, but also in terms of genetic content.

The famous sheep Dolly became the first mammal to be cloned in 1966. It was obtained by transplanting the nucleus of a somatic cell into the cytoplasm of the egg. Dolly was a genetic copy of the sheep who donor the cell nucleus. Under natural conditions, an individual is formed from one fertilized egg, having received half of the genetic material from two parents. However, during cloning, the genetic material was taken from the cell of one individual. First, the nucleus, which contains the DNA itself, was removed from the zygote. Then the nucleus was removed from the cell adult sheep and implanted it into that nucleated zygote, and then it was transplanted into the uterus of an adult and given the opportunity to grow and develop.

However, not all cloning attempts were successful. In parallel with Dolly's cloning, a DNA replacement experiment was carried out on 273 other eggs. But only in one case was a living adult animal able to fully develop and grow. After Dolly, scientists tried to clone other mammal species.

One of the types of genetic engineering is genome editing.

The CRISPR/Cas tool is based on an element of the immune defense system of bacteria, which scientists have adapted to introduce any changes to the DNA of animals or plants.

CRISPR/Cas is one of the biotechnological methods for manipulating individual genes in cells. There are a huge number of applications for this technology. CRISPR/Cas allows researchers to figure out the function of different genes. To do this, you simply need to cut out the gene of interest from the DNA and study which body functions were affected.

Some practical applications systems:

1. Agriculture. CRISPR/Cas systems can be used to improve crops. Namely, to make them more tasty and nutritious, as well as heat-resistant. It is possible to endow plants with other properties: for example, cut out an allergen gene from nuts (peanuts or hazelnuts).
2. Medicine, hereditary diseases. Scientists have a goal of using CRISPR/Cas to remove mutations from the human genome that can cause diseases such as sickle cell anemia, etc. In theory, using CRISPR/Cas it is possible to stop the development of HIV.
3. Gene drive. CRISPR/Cas can change not only the genome of an individual animal or plant, but also the gene pool of a species. This concept is known as "gene drive". Every living organism passes half of its genes to its offspring. But using CRISPR/Cas can increase the likelihood of gene transfer by up to 100%. This is important so that the desired trait spreads faster throughout the population.

Swiss scientists have significantly improved and modernized the CRISPR/Cas genome editing method, thereby expanding its capabilities. However, scientists could only modify one gene at a time using the CRISPR/Cas system. But now researchers at ETH Zurich have developed a method that can simultaneously modify 25 genes in a cell.

For the newest technique, specialists used the Cas12a enzyme. Geneticists have successfully cloned monkeys for the first time in history. "Popular Mechanics";

Nikolenko S. (2012). Genomics: Problem Statement and Sequencing Methods. "Postscience".

A molecular biologist is a medical researcher whose mission is, no less than, to save humanity from dangerous diseases. Among such diseases, for example, oncology, which today has become one of the main causes of mortality in the world, only slightly inferior to the leader - cardiovascular diseases. New methods for early diagnosis of oncology, prevention and treatment of cancer are a priority modern medicine. Molecular biologists in oncology develop antibodies and recombinant (genetically engineered) proteins for early diagnosis or targeted drug delivery in the body. Specialists in this field use the most modern achievements of science and technology to create new organisms and organic substances for their further use in research and clinical activities. Among the methods that molecular biologists use are cloning, transfection, infection, polymerase chain reaction, gene sequencing, and others. One of the companies interested in molecular biologists in Russia is PrimeBioMed LLC. The organization is engaged in the production of antibody reagents for the diagnosis of cancer. Such antibodies are mainly used to determine the type of tumor, its origin and malignancy, that is, the ability to metastasize (spread to other parts of the body). Antibodies are applied to thin sections of the tissue being examined, after which they bind in cells to certain proteins - markers that are present in tumor cells but absent in healthy ones and vice versa. Depending on the results of the study, further treatment is prescribed. Among PrimeBioMed's clients are not only medical, but also scientific institutions, since antibodies can also be used to solve research problems. In such cases, unique antibodies can be produced that can bind to the protein of interest, under specific task by special order. Another promising area of ​​research for the company is targeted delivery of drugs in the body. In this case, antibodies are used as transport: with their help, drugs are delivered directly to the affected organs. Thus, the treatment becomes more effective and has fewer negative consequences for the body than, for example, chemotherapy, which affects not only cancer cells, but also other cells. The profession of a molecular biologist is expected to become increasingly in demand in the coming decades: with an increase in average duration Throughout human life, the number of cancer diseases will increase. Early diagnosis of tumors and innovative treatment methods using the obtained molecular biologists substances will save lives and improve their quality for a huge number of people.