The objects of biotechnology are viruses, bacteria, fungi - micromycetes and macromycetes, protozoan organisms, cells (tissues) of plants, animals and humans, some biogenic and functionally similar substances (for example, enzymes, prostaglandins, lectins, nucleic acids, etc.). Therefore, biotechnology objects can be represented by organized particles (viruses), cells (tissues) or their metabolites (primary, secondary). Even when using a biomolecule as an object of biotechnology, its initial biosynthesis is carried out in most cases by the corresponding cells. In this regard, we can say that the objects of biotechnology belong either to microbes, or to plant and animal organisms. In turn, the body can be figuratively characterized as a system of economical, complex, compact, self-regulating and, therefore, targeted biochemical production, which proceeds stably and actively while optimally maintaining all the necessary parameters. From this definition it follows that viruses are not organisms, but according to the content of the molecules of heredity, adaptability, variability and some other properties, they belong to representatives of wildlife.

As can be seen from the diagram, biotechnology objects are extremely diverse, their range extends from organized particles (viruses) to humans.

Viruses occupy a position between living and nonliving nature, they do not have a nucleus, although there is hereditary nuclear material - ribonucleic acid (RNA) or deoxyribonucleic acid (DNA).

Unlike microbes, cellular organization of RNA and DNA in viral particles is never detected together.

Currently, most of the biotechnological objects are microbes belonging to the three kingdoms (nuclear-free, pre-nuclear, nuclear) and five kingdoms (viruses, bacteria, fungi, plants and animals). Moreover, the first two kingdoms consist exclusively of microbes, while the third - mainly from plants and animals.

In the first half of the XIX century. one of the most basic generalizations of biology was made - the cellular theory (M. Schleiden, T. Schwann, R. Virchow), which became generally recognized. It turned out to be the foundation of science - cytology (from the Greek. Kitos - cavity). Of all the objects of biotechnology, only viruses, viroids and biomolecules do not have cellular organization. However, viruses, being in cells, behave like living beings - they replicate (“multiply”) and their genetic material functions mainly according to the general laws inherent in cells of any origin. With the improvement of methods and techniques of cytological studies, scientists penetrate deeper into the essence of organized particles and cells, and as a result of such penetration it is possible to substantiate the belonging of all living beings to three kingdoms: Acaryotae - non-nuclear, Procaryotae - pre-nuclear and Eucaryotae - nuclear (from Greek a - no, pro is up, she is good, full, karyon is the core). The first includes organized particles - viruses and viroids, the second - bacteria, the third - all other organisms (fungi, algae, plants, animals).

Despite the fact that representatives of all the supra-kingdoms contain genetic material, various acariotes are devoid of any one type of nucleic acid RNA or DNA. They are not able to function (including replicate) outside a living cell, and, therefore, it is legitimate to call them nuclear-free.

The cells of fungi, algae, plants and animals have a real nucleus that is delimited from the cytoplasm and therefore they are referred to as eukaryotes.

The classification of prokaryotes and eukaryotes is based on numerous structural differences, the main ones are as follows: 1) the presence or absence of a nucleus containing chromosomal DNA; 2) the structure and chemical composition of the cell wall; and 3) the presence or absence of subcellular cytoplasmic organelles. In a prokaryotic cell, such as a bacterial one, the chromosomal DNA is located directly in the cytoplasm, the cell is surrounded by a rigid cell wall, which often includes peptidoglycan, but not chitin or cellulose; there are no subcellular cytoplasmic organelles in the cell. In a eukaryotic cell there is a nucleus separated from the cytoplasm by a nuclear membrane; chromosomal DNA is in the nucleus; the cell wall, if present, may contain chitin or cellulose, but not peptidoglycan; the cytoplasm contains various subcellular organelles (mitochondria, Golgi apparatus, chloroplast in plant cells) (Fig. 1).

Fig. 1. Scheme of prokaryotic bacterial cells (A) and eukaryotic animal cells   (B)

Nucleic acids are substances of the heredity of viruses. According to the type of nucleic acid, they are divided into RNA-containing viruses and DNA-containing viruses. The first includes all plant viruses, the second - the majority of bacteriophages, a number of human and animal viruses (adenoviruses, herpes viruses, smallpox vaccines, etc.).

A protein is structured around a viral nucleic acid (genome) in the form of a shell and is called a capsid. The shape of the virion is determined by its capsid. Together with nucleic acid, the capsid forms a nucleocapsid.

A sample list of viruses includes 17 families of vertebrate viruses and 7 families of invertebrate viruses, 10 families of bacteria viruses. Describes 20 genera of plant viruses and 5 genera of fungal viruses. The classification schemes of viruses are not yet fully established, and they also open up viruses that are new to science (for example, the viruses of Ebola and human immunodeficiency). Representatives of DNA-containing viruses are molluscum contagiosum, smallpox, herpes viruses, most bacterial phages; RNA-containing viruses are plant viruses, human influenza viruses, rabies, poliomyelitis, etc.

Viroids.   By their molecular structure, viroids are single-stranded, covalently closed, ring RNA molecules lacking capsids. The number of nucleotides in such RNA is in the range of 240-400. The shape of the viroid can be linear and ring-shaped, they are able to take a hairpin, quasi-double-chain conformation (from lat. Quasi - supposedly, as if almost, close; conformatio - shape, location). Each type of viroid contains a unique, unique to it, special type of low molecular weight RNA. The size of viroids is within 15 nm. In sensitive cells of host plants, they concentrate in the nucleus, associating with the nucleolus in the form of a protein-nucleic complex, and replicate autonomously as a whole with the help of previous or activated host enzymes. Viroids are not broadcast. This is confirmed by their structural similarity between themselves and the absence of initiator codons in a number of viroids. At the same time, replication occurs due to transcription of viroid RNA sequences from RNA matrices with the participation of RNA polymerases.

Bacteria are creatures of the cell organization in which the nuclear material is not separated from the cytoplasm by elementary membranes and is not associated with any basic proteins. The cytoplasm in them with irregularly scattered ribosomes is motionless, the cells do not have the ability to endo- and exocytosis. Most bacteria are unicellular, their smallest diameter is 0.2-10.0 microns.

All bacteria make up the united kingdom of Bacteria, although some of them - archaeobacteria (Archaeobacteria) are noticeably different from others called eubacteria (Eubacteria). Obviously, archaeobacteria are more ancient representatives of prokaryotes than eubacteria. They live in environments with extreme conditions - high concentrations of inorganic salts, elevated temperatures, carbon monoxide and dioxide - as the only sources of carbon. Archobacteria include halobacteria, thermoacidophilic bacteria and methane-forming, or methanogenic bacteria.

Phototrophic bacteria are oxygen cyanobacteria, anoxigenic purple and green bacteria; chemotrophic - gram-positive and gram-negative bacteria and bacilli, myxobacteria, stalk and budding bacteria, vibrios, spirillas, spirochetes, actinomycetes, corynebacteria, mycobacteria, rickettsia, chlamydia, mycoplasma and spiroplasm.

Bacterium   Esherichia   coli   - one of the best-studied organisms. In recent years, it has been possible to obtain comprehensive information about its genetics, molecular biology, biochemistry, physiology and general biology. This is a gram-negative non-pathogenic bacillus less than 1 micron in length. Its habitat is the human intestines, but it can also be sown from soil and water. Due to its ability to multiply by simple division on media containing only Na +, K +, Mg 2+, Ca 2+, NH 4 +, Cl ~, НР0 4 2 ~ and S0 4 2 ~ ions, trace elements and a carbon source (e.g. glucose ),   E.   coli   became a favorite subject of scientific research. Under cultivation   E.   coli   on enriched liquid nutrient media containing amino acids, vitamins, salts, trace elements and a carbon source, the generation time (i.e., the time between the formation of a bacterium and its division) in the logarithmic phase of growth at a temperature of 37 ° C is approximately 22 minutes.

For every living organism, there is a certain temperature range that is optimal for its growth and reproduction. At too high temperatures, protein denaturation and destruction of other important cellular components occurs, which leads to cell death. At low temperatures, biological processes significantly slow down or stop completely due to structural changes that undergo protein molecules. Based on the temperature regime that certain microorganisms prefer, they can be divided into thermophiles (from 45 to 90 ° C and above), mesophiles (from 10 to 47 ° C) and psychrophiles, or psychrotrophs (from -5 to 35 ° C) ) Microorganisms that actively reproduce only in a certain temperature range can be a useful tool for solving various biotechnological problems. For example, thermophiles often serve as a source of genes encoding thermostable enzymes that are used in industrial or laboratory processes, and genetically modified psychrotrophs are used for biodegradation of toxic waste contained in soil and water at low temperatures.

  E.   coli   can be cultivated both under aerobic (in the presence of oxygen) and anaerobic (without oxygen) conditions. However, for optimal production of recombinant proteins   E.   coli   and other microorganisms are usually grown under aerobic conditions. If the purpose of the cultivation of bacteria in the laboratory is the synthesis and isolation of a specific protein, then the cultures are grown on complex liquid nutrient media in flasks. To maintain the desired temperature and ensure sufficient aeration of the culture medium, the flasks are placed in a water bath or a temperature-controlled room and shaken continuously. Such aeration is sufficient for cell reproduction, but not always for protein synthesis. The growth of cell mass and protein production are limited not by the content of carbon or nitrogen sources in the nutrient medium, but by the content of dissolved oxygen: at 20 ° С it is equal to approximately nine ppm. This becomes especially important in the industrial production of recombinant proteins using microorganisms. To ensure optimal conditions for maximum protein production, special fermenters are designed and aeration systems are created.

Besides   E.   coli, in molecular biotechnology, many other microorganisms are used. They can be divided into two groups: microorganisms as sources of specific genes and microorganisms created by genetic engineering methods to solve certain problems. Specific genes include, for example, a gene encoding a thermostable DNA polymerase, which is used in the widely used polymerase chain reaction (PCR). This gene was isolated from thermophilic bacteria and cloned into   E.   coli. The second group of microorganisms include, for example, various strains   Corynebacterium   glutamicumthat have been genetically modified to increase the production of industrially important amino acids.

The biotechnological functions of fungi are diverse. They are used to obtain products such as:

Antibiotics (penicillas, streptomycetes, cephalosporins);

Gibberellins and cytokinins (Physarium and Botritis);

· Carotenoids (for example, astaxanthin, which gives the pulp of salmon fish a red-orange hue, is produced by Rhaffia rhodozima, which is added to the feed at the fish factories);

Protein (Candida, Saccharomyces lipolitica);

Cheeses such as Roquefort and Camembert (penicillas);

Soy sauce (Aspergillus oryzae).

Mushrooms include actinomycetes, yeast, and mold. True actinomycetes are strict aerobes, they are gram-positive and do not form a dispute. The most representative in this group is the genus Streptomyces, some species of which produce widely used antibiotics. When growing on solid media, actinomycetes form a very thin mycelium with aerial hyphae that differentiate into chains of conidiospores. Each conidiospore is capable of forming a microcolony.

Antibiotics are also produced by another type of actinomycetes, Micromonospora, whose colonies lack air hyphae and form conidiospores directly on the mycelium.

Of the 500 known yeast species, the first people learned to use Saccharomyces cerevisiae, this species is most intensively cultivated. Yeast   Saccharomyces   cerevisiae   Are non-pathogenic unicellular microorganisms with a cell diameter of approximately 5 μm, which in many respects are a eukaryotic analogue   E.   coli. Their genetics, molecular biology and metabolism are studied in detail.   S.   cere   visiae   reproduce by budding and grow well in the same simple environment as   E.   coli. Their ability to convert sugar to ethanol and carbon dioxide has long been used to make alcoholic beverages and bread. Yeast   S.   cerevisiae are also of great scientific interest. In particular, they are the most convenient model for the study of other eukaryotes, including humans, since many genes are responsible for the regulation of cell division   S.   cerevisiaeare similar to those in humans. This discovery contributed to the identification and characterization of human genes responsible for the development of neoplasms. The widely used genetic system of yeast (artificial chromosome) is an indispensable participant in all studies on the study of human DNA. In 1996, the complete nucleotide sequence of the entire set of chromosomes was determined   S.   cerevisiae, which further increased the value of this microorganism for scientific research.

A eukaryotic protein synthesized by a bacterial cell often has to undergo enzymatic modification by attaching low molecular weight compounds to the protein molecule - in many cases this is necessary for the protein to function properly. Unfortunately,   E.   coli   and other prokaryotes are not able to carry out these modifications, therefore, to obtain complete eukaryotic proteins,   S.   cerevisiaeas well as other types of yeast:   Kluyveromyces   lactis,   Saccharomyces   diastaticus,   Schizisaccharomyces   pombe,   Yarrowia   lipolytica,   Pichia   pastoris,   Hansenula   polymo. The most effective producers of complete recombinant proteins are   P.   pastoris   and N.   polymo.

Lactose Fermenting Yeast   Kluyveromyces fragiliswhich is used to produce alcohol from whey.   Saccharomycopsis lipolytica   degrades hydrocarbons and is used to obtain protein mass. All three species belong to the class of ascomycetes. Other useful species belong to the class of deuteromycetes (imperfect mushrooms), since they do not reproduce sexually, but by budding.   Candida utilis   grows in sulfite wastewater (paper industry waste).   Trichosporon cutaneum,   oxidizing numerous organic compounds, including some toxic ones (such as phenol), plays an important role in aerobic wastewater treatment systems.   Phaffia rhodozyma   synthesizes astaxanthin - a carotenoid that gives the pulp of trout and salmon grown on farms, a characteristic orange or pinkish color. Industrial yeast usually does not reproduce sexually, does not form spores and is polyploid. The latter explains their strength and ability to adapt to changes in the culture medium (normal cell nucleus   S.cerevisiae   contains 17 or 34 chromosomes, i.e. cells are either haploid or diploid).

Mildew causes numerous transformations in solid media that occur before fermentation. Their presence explains the hydrolysis of rice starch in the production of sake and the hydrolysis of soybeans, rice and malt when receiving food used in Asian countries. Mold-based Fermented Foods   Rhizopus oligosporus   soybeans or wheat contain 5-7 times more vitamins such as riboflavin, nicotinic acid) and are distinguished by several times higher protein content. Molds also produce enzymes used in industry (amylases, pectinases, etc.), organic acids and antibiotics. They are also used in the production of cheeses, for example, Camembert and Roquefort.

Artificial mushroom cultivation is capable of making another, equally important contribution to the provision of food to the growing population of the globe. People eat mushrooms since ancient times. Therefore, to make mushrooms the same managed crop as cereals, vegetables, fruits, has long been an urgent task. Wood-destroying mushrooms are most easily amenable to artificial cultivation. This is due to the peculiarities of their biology, which we have become aware and understand only now. Their ability to grow and bear fruit easily has been used since ancient times.

Artificial breeding of wood-destroying fungi is quite widespread. The mycelium of edible fungi can be grown on liquid media, for example, on whey, etc., in special fermenters, in the so-called deep culture.

The simplest are among the non-traditional objects of biotechnology. Until recently, they were used only as a component of activated sludge in the biological treatment of wastewater. Currently, they have attracted the attention of researchers as producers of biologically active substances.

In this capacity, it is more rational to use free-living protozoa that have diverse biosynthetic capabilities and are therefore widely distributed in nature.

A special ecological niche is occupied by protozoa living in the rumen of ruminants. They possess the enzyme cellulase, which promotes the decomposition of fiber in the stomach of ruminants. Protozoa can be the source of this valuable enzyme. The causative agent of South American trypanosomiasis is   Trypanosoma (Schizotrypanum cruzi) became the first producer of the anticancer drug crucine (USSR) and its analogue trypanose (France). Studying the mechanism of action of these drugs, scientists came to the conclusion that these drugs have a cytotoxic effect in direct contact with the tumor and inhibit it indirectly by stimulating the reticuloendothelial system. It turned out that the inhibitory effect is associated with fatty acid fractions. A characteristic feature of these organisms is the high content of unsaturated fatty acids, which is 70-80% in trypanosomids, and in   Astasia longa   (free-living flagellate) - 60% of the sum of all fatty acids. In flagellates, phospholipids and polyunsaturated fatty acids have the same composition and structure as in humans and animals. In the world of microbes, polyunsaturated fatty acids are not synthesized, and multicellular animals or plants represent a more limited raw material base than protozoa, whose cultures can be obtained by biotechnological methods regardless of the season or climatic conditions.

Since the lipid metabolism of protozoa has relative lability, the ways of its regulation were studied. Application to the simplest methods of microbiology generally accepted in microbiology of increasing lipid biosynthesis by reducing the nitrogen source content in the medium and increasing the carbon source content led to a sharp inhibition or stop of the growth of cultures. To create conditions for targeted lipid biosynthesis, precursors and stimulators of lipid biosynthesis: malonate, citrate, succinate, cytidine nucleotides in combination with a specific aeration regimen were added to the medium for cultivation of flagellates.

Another group of biologically active substances of protozoa is polysaccharides. The variety of polysaccharides synthesized by protozoa is quite large. Of particular interest is paramilon, characteristic of euglenoid flagella. Representatives of the birth   Astasia and Euglena   capable of super-synthesis of paramilon, which constitutes over 50% of the dry residue of cells. This polysaccharide is being studied as a stimulator of the mammalian immune system. Paramilon isolated from A. longa is practically non-toxic. The pronounced immunomodulatory effect and low toxicity of this drug are the prerequisites for its in-depth studies in combination with direct antitumor drugs, radiotherapy and other adjuvants.

Currently, the world attaches great importance to the production of glucans, not only for medical purposes, but also for the food and textile industries. Until now, glucans have been obtained from cultures of bacteria or algae. Euglenides are one of the most promising sources of this substance. The structural polysaccharides that make up the cell membranes of protozoa are heteropolysaccharides containing glucose, mannose, xylose, arabinose, ribose, galactose, rhamnose, fructose, glucosamine. The most characteristic heteropolysaccharides are arabinogalactans, D-galacto-D-mannan, phosphanoglucans and others.

Protozoa biomass contains up to 50% protein. Its high biological value lies in the fact that it contains all the essential amino acids, and the content of free amino acids is an order of magnitude higher than in the biomass of microalgae, bacteria and meat. This indicates the wide possibilities of using free-living protozoa as a source of feed protein.

Algae are mainly used to produce protein. In this respect, cultures of unicellular algae, in particular, highly productive strains of the genus, are also very promising.   Chlorella and Scenedesmus. After appropriate treatment, their biomass is used as an additive in livestock diets, as well as for food purposes.

Unicellular algae are grown in mild warm climates (Central Asia, Crimea) in open pools with a special nutrient medium. For example, during the warm period of the year (6-8 months), you can get 50-60 tons of chlorella biomass per 1 ha, while one of the most highly productive herbs - alfalfa gives only 15-20 tons of crop from the same area.

Chlorella contains about 50% protein, and alfalfa - only 18%. In general, per 1 ha, chlorella forms 20-30 tons of pure protein, and alfalfa - 2-3.5 tons. In addition, chlorella contains 40% carbohydrates, 7-10% fats, vitamins A (20 times more), B2, K, PP and many trace elements. By varying the composition of the nutrient medium, biosynthesis in the chlorella cells can be shifted towards the accumulation of either proteins or carbohydrates, as well as activation of the formation of certain vitamins.

At least 100 species of macrophyte algae are consumed in food both in Europe and America, and especially in the East. A lot of various dishes are prepared from them, including diet, salads, seasonings. They are served in the form of candied pieces, a kind of candy, they make jam, make jelly, additives to the dough and much more. In the store you can buy canned seaweed - kelp of the Far Eastern or Northern Seas. It is canned with meat, fish, vegetables, rice, used in the preparation of soups, etc. It, along with the microalgae chlorella, is the most popular edible and fodder algae.

Other edible macrophytic algae are also known - ulva, from which various green salads are made, as well as alaria, porphyry, paternity, chondrus, undaria, and others. In Japan, products obtained from kelp are called “kombu”, and in order to make them tasty cook, there are more than a dozen ways.

In a number of countries, algae is used as a very useful vitamin supplement to feed for farm animals. They are added to hay or given as an independent feed for cows, horses, sheep, goats, poultry in France, Scotland, Sweden, Norway, Iceland, Japan, America, Denmark and in our North. The biomass of the grown microalgae (chlorella, scenesdesmus, dunaliella, etc.) is also fed to animals as an additive.

Green algae protein hydrolysates   Scenedesmus   used in medicine and cosmetic industry. In Israel, experimental plants are conducting experiments with green unicellular algae   Dunaliella bardaw   il, which synthesizes glycerol. This algae belongs to the class of equal-flagged and similar to chlamydomonas.   Dunadiella can grow and multiply in an environment with a wide range of salt content: in the water of the oceans, and in the almost saturated salt solutions of the Dead Sea. It accumulates free glycerol to counteract the adverse effects of high salt concentrations in the environment where it grows. Under optimal conditions and a high salt content, glycerol accounts for up to 85% of the dry cell mass. For growth, these algae require: seawater, carbon dioxide and sunlight. After processing, these algae can be used as animal feed, since they do not have the indigestible cell membrane inherent in other algae. They also contain significant amounts of β-carotene. Thus, cultivating this algae, you can get glycerol, pigment and protein, which is very promising from an economic point of view.

Along with feed, algae have long been used in agriculture as fertilizers. Biomass enriches the soil with phosphorus, potassium, iodine and a significant amount of trace elements, also replenishes its bacterial, including nitrogen-fixing, microflora. At the same time, algae decompose faster in the soil than manure fertilizers, and do not clog it with weed seeds, larvae of harmful insects, and spores of phytopathogenic fungi.

One of the most valuable products obtained from red algae is agar, a polysaccharide present in their shells and consisting of agarose and agaropectin. Its amount reaches 30-40% of the weight of algae (algae laurence and gracillaria, gelidium). Algae is the only source of agar, agaroids, carrageenin, alginates.

Brown algae are the only source of one of the most valuable algae substances - salts of alginic acid, alginates. Alginic acid is a linear heteropolysaccharide built from linked residues (3-D-mannuronic and α-L-hyaluronic acids.

Alginates are used in the national economy. This is the manufacture of high-quality lubricants for rubbing machine parts, medical and perfume ointments and creams, synthetic fibers and plastics, weather-resistant coatings, fabrics that do not fade over time, production of silk, extremely strong adhesives, building materials, high-quality food products - fruit juices, canned food, ice cream, solution stabilizers, fuel briquetting, foundry and much more. Sodium alginate is able to absorb up to 300 weight units of water, while forming viscous solutions.

Brown algae is also rich in a very useful compound - six-atom alcohol mannitol, which is used in the food industry, pharmaceuticals, in the manufacture of paper, paints, explosives, etc. Brown algae is planned to be used for biogas production in the near future. Callus cultures of macrophytic algae can be used further in various directions. In case they are obtained from agarophytes, agar can be directly obtained from them. Callus cultures of edible macrophytic algae, for example, laminaria, can in the future be used to produce protein that is directly consumed in food and in nutritional supplements, as well as in feed for farm animals.

Higher plants (about 300,000 species) are differentiated multicellular, mainly terrestrial organisms. In the process of differentiation and specialization, plant cells were grouped into tissues (simple - from the same type of cells, and complex - from different types of cells). Tissues, depending on the function, are divided into educational, or meristemic (from the Greek. Meristos - divisible), integumentary, conductive, mechanical, basic, secretory (excretory). Of all the tissues, only meristematic tissues are capable of division, and due to them all other tissues are formed. It is important to obtain cells, which then must be included in the biotechnological process.

Meristem cells that linger at the embryonic stage of development throughout the life of the plant are called initial, others gradually differentiate and turn into cells of various permanent tissues - the final cells. Any kind of plant can produce, under appropriate conditions, an unorganized mass of dividing cells - callus   (from lat. callus - corn), especially with the inducing effect of plant hormones. Mass production of calluses with further regeneration of shoots is suitable for large-scale plant production. In general, callus is the main type of plant cell cultured on a nutrient medium. Callus tissue from any plant can be recultivated for a long time. In this case, the initial plants (including meristematic ones) are dedifferentiated and despecialized, but are induced to divide, forming a primary callus.

In addition to growing calluses, it is possible to cultivate the cells of some plants in suspension cultures.

Protoplasts of plant cells also appear to be important bioobjects. The methods for their preparation are fundamentally similar to the methods for producing bacterial and fungal protoplasts.

In addition to the culture of plant cells, water azoll fern is used. It is valued as an organic nitrogen fertilizer, as it grows in close symbiosis with the blue-green alaba algae. This allows the symbiotic anaben-azoll organism to accumulate a lot of nitrogen in the vegetative mass. Anabenu-azolla is grown in rice fields before sowing rice, which reduces the amount of fertilizer applied.

Representatives of the duckweed family   (Lemnaceae)   - the smallest and simplest in structure flowering plants, the size of which rarely exceeds 1 cm. Raskovye - free-living aquatic floating plants. The vegetative body resembles a leaf or thallus of lower plants; therefore, until the beginning of the 18th century, duckweed was classified as thallus.

Raskaskovye (   Lemna minor, L. trisulca, Wolfia, Spirodela polyrhiza) serve as animal feed, for ducks and other waterfowl, fish, muskrat. They are used both fresh and dry as a valuable protein feed for pigs and poultry. Cossacks contain a lot of protein (up to 45% of dry weight). 45% carbohydrates, 5% fat and the rest is fiber, etc. They are highly productive, unpretentious in culture, well purify water and enrich it with oxygen. This makes duckweed a valuable object for morphogenetic, physiological and biochemical studies.

As objects of biotechnology, the animals themselves and animal cell cultures can be used.

Despite all the differences between the types of eukaryotes, methodological approaches to the cultivation of insect, plant, and mammalian cells have much in common. First, they take a small piece of the tissue of this organism and treat it with proteolytic enzymes that break down the proteins of the intercellular material (when working with plant cells, special enzymes that destroy the cell wall are added). The released cells are placed in a complex nutrient medium containing amino acids, antibiotics, vitamins, salts, glucose and growth factors. Under these conditions, cells divide until a cell monolayer forms on the walls of the culture reservoir. If after this you do not transfer cells to containers with fresh nutrient medium, then growth will stop. Usually it is possible to transfer (transplant, subculture) and maintain up to 50-100 cell generations of the initial (primary) cell culture, then the cells begin to lose their ability to divide and die. Cultured cells retain some properties of the original cellular material, so they can be used to study the biochemical properties of various tissues.

Often, some cells of transplanted primary cell cultures undergo genetic changes, as a result of which their growth is accelerated. Cell cultures, which at the same time acquire selective advantages, are capable of unlimited growth in vitro and are called resistant cell lines. Some cell lines retain the basic biochemical properties of the original cells, while others do not. Most cells capable of unlimited growth have significant chromosome changes, in particular, an increase in the number of some chromosomes and a loss of others. In molecular biotechnology, resistant animal cell lines are used to propagate viruses and to detect proteins that are encoded by cloned DNA sequences. In addition, they are used for large-scale production of vaccines and recombinant proteins.

For the implementation of biotechnological processes, important parameters of bioobjects are: purity, rate of cell reproduction and reproduction of viral particles, activity and stability of biomolecules or biosystems.

It should be borne in mind that when creating favorable conditions for a selected bio-object of biotechnology, these same conditions may turn out to be favorable, for example, for microbes - contaminants, or pollutants. Representatives of contaminating microflora are viruses, bacteria and fungi found in the cultures of plant or animal cells. In these cases, microbial contaminants act as pests of production in biotechnology. When using enzymes as biocatalysts, it becomes necessary to protect them in an isolated or immobilized state from the destruction of banal saprophytic (non-pathogenic) microflora, which can penetrate the sphere of the biotechnological process from the outside due to the non-sterility of the system.

Activity and stability in the active state of biological objects are one of the most important indicators of their suitability for long-term use in biotechnology.

Thus, regardless of the systematic position of the bioobject, in practice they use either natural organized particles (phages, viruses) and cells with natural genetic information, or cells with artificially set genetic information, that is, in any case, they use cells, be it a microorganism, plant, animal or human. For example, we can name the process of obtaining polio virus on a cell culture of monkey kidneys in order to create a vaccine against this dangerous disease. Although we are interested here in the accumulation of the virus, its reproduction takes place in the cells of the animal organism. Another example is with enzymes that will be used in an immobilized state. The source of enzymes are also isolated cells or their specialized associations in the form of tissues, from which the necessary biocatalysts are isolated.

One of the basic concepts in biotechnology is the concept of "biosystem". The generalized characteristics of a biological system (living system) can be reduced to three characteristics.

• 1. Living systems are heterogeneous open systems that exchange substances and energy with the environment.
• 2. These systems are self-governing, self-regulatory, capable of exchanging information with the environment to maintain its structure and manage metabolic processes.
• 3. Living systems are self-reproducing.

Unicellular organisms should be named as biological systems, objects that biotechnology uses, among them there are groups of acariotes (viruses), prokaryotes (bacteria, blue-green algae) and eukaryotes (fungi, protozoal, algae). Their sizes range from nanometers (viruses, bacteriophages) to millimeters and centimeters (giant algae). In addition to the above, cells and tissues of plants, animals and humans, substances of biological origin (for example, enzymes, prostaglandins, pectins, nucleic acids), and molecules are used as objects of biotechnology.

The selection of these objects is due to the following points:

• 1. Cells are a kind of “biofactories” that produce a variety of valuable products in the process of life: proteins, fats, carbohydrates, vitamins. nuclein. acids, antibiotics, antibodies, enzymes, alcohols, etc. Many of these products, which are extremely necessary for a person, are not yet available for production using non-biotechnological methods due to scarcity or high cost of raw materials or complexity of technological processes.
• 2. Cells reproduce extremely quickly. So, the bacterial cell divides every 20-60 minutes, the yeast - after 1.5-2 hours, the animal - after 24 hours, which allows for a relatively short time to artificially grow on relatively cheap and non-deficient nutrient media in industrial. scales of huge amounts of biomass of microbial, animal and plant cells. In the process of life during their cultivation, a large amount of valuable products enters the environment.
• 3. The biosynthesis of complex substances, such as proteins, antibiotics, antigens, etc. is much more affordable and cheaper than chemical synthesis. As raw materials for biosynthesis use wastes from agricultural, fish products, food industry, plant materials (yeast, wood, etc.)
• 4. The possibility of carrying out a biotechnological process on an industrial scale, i.e. availability of appropriate technological equipment, availability of raw materials, processing technologies.

The methods used in biotechnology are determined by two levels: cellular and molecular. Both that and another are defined by bioobjects.

In the first case, they deal with bacterial cells (to obtain vaccines), actinomycetes (when receiving antibiotics), micromycetes (when receiving citric acid), animal cells (when making antiviral vaccines), human cells (when making interferon), etc.

In the second case, they deal with molecules, for example, nucleic acids. However, in the final stage, the molecular level is transformed into a cellular level.

When choosing a biological object in all cases, the principle of manufacturability must be observed. So, if during numerous cultivation cycles the properties of a biological object are not preserved or undergo changes, then this object should be recognized as non-technological, i.e. unacceptable for the following technological development following the stage of laboratory research.

Scientists devote much attention to targeted creation new biological objects that do not exist in nature.   First of all, it should be noted the creation of new cells of microorganisms, plants, animals by genetic engineering methods. The creation of new biological objects, of course, contributes to the improvement of the legal protection of inventions in the field of genetic engineering and biotechnology in general. Formed a direction engaged in the construction of artificial cells. Currently, there are methods to obtain artificial cells using various synthetic and biological materials, for example, an artificial cell membrane with a given permeability and surface properties. Some materials can be enclosed inside such cells: enzyme systems, cell extracts, antibodies, hormones, biological cells, etc.

For example, the use of artificial cells has given positive results in the production of interferons, immunosorbents.

Also, scientists are using and anaerobic organisms.   Anaerobic processes attract the attention of researchers due to a lack of energy and the possibility of obtaining biogas.

Anaerobic microorganisms are successfully used for processing waste (plant biomass, food industry waste, household waste, etc.) and sewage (domestic and industrial waste, manure) into biogas.

As follows from the foregoing, in biotechnological processes it is possible to use a number of biological objects characterized by different levels of complexity: cellular, subcellular, molecular. The approach to creating the entire biotechnological system as a whole depends on the characteristics of a particular biological object.

Microscopic mushrooms as an object of biotechnology

This lesson completes the study of the topic "Fundamentals of Biotechnology", which is an independent section of the topic "Fundamentals of Selection" in the 11th grade with an in-depth study of biology.

The purpose of the lesson:students learning about the use of microscopic mushrooms in the food, pharmaceutical industry, agriculture, in the disposal of household waste, as an object of biotechnological research aimed at optimizing existing and creating new industries.

Equipment:term signs attached to a metal board with magnets, the table “Microbial methods of recycling” (on each desk), tests to control knowledge, costumes for dramatization.

DURING THE CLASSES

Revitalization of cognitive activity

Recovery in memory of previously studied material.

- What topic are we studying? (Basics of biotechnology.)
   - What is biotechnology? (The use of living organisms and biological processes in production.)
   - What objects of biotechnology did we meet in previous lessons? (With bacteria.)
   - What is the name of the science that studies mushrooms? (Mycology.)

Learning New Material

The teacher formulates the main objectives of the lesson.

B. Viewing the staging “Microscopic Mushrooms” (see Biology, No. 5/1997).

B. Discussion of the play. (In the course of the discussion, a diagram is drawn on the board.)

1. Yeast as the most studied object of biotechnological research

(performance of the student who played in the play Yeast Cell)

Yeast is a group of fungi that do not have a typical mycelium and exist as separate budding or dividing cells.
   About 500 types of yeast are known. All yeast are heterotrophs with an oxidative (respiration) or fermentation (fermentation) type of metabolism. Yeast synthesizes proteins, lipids, extracellular polysaccharides, B vitamins. Causes diseases: thrush (cryptococcosis, candidiasis) and other mycoses.
   Human use: brewing, winemaking, alcohol industry, bakery, microbiological industry (feed protein, enzymes), as well as an object of research in bioenergy, radiobiology, genetics.
   Most of the species used by humans belong to the genus saccharomycetes ( Saccharomyces) from the class of ascomycetes ( Ascomycota), which actively ferment simple carbohydrates to ethyl alcohol. Alcoholic fermentation was first studied in detail by Louis Pasteur.

The scheme of oxidation of carbohydrates to ethanol:

sugar ---\u003e pyruvate ---\u003e CO 2 + acetaldehyde ---\u003e ethanol.

The genetics of baker's yeast has been studied in detail. S.cerevisiae. By genetic engineering, the genes responsible for the synthesis of hormones and other valuable compounds are inserted into and cloned (“propagated” during replication of chromosomal DNA) into the chromosomes of yeast cells.
   The properties of yeast that are valuable for biotechnology: they grow rapidly, are safe for humans, and grow on a cheap environment (paraffin, molasses, methyl alcohol). The disadvantage is that it is difficult to obtain intracellular products, because cells are coated with a very strong membrane. The most commonly used method for producing intracellular compounds is autolysis, i.e. cell destruction by its own enzymes.

Bakery.Earlier in bread baking, yeast dough was widely used. It is now widely used for baking rye bread, as well as in the household. To obtain such a test, use a dough - a small portion of the dough left from the previous batch or kneaded in advance to the main batch. Yeast and lactic acid bacteria are contained and propagated in dough, giving brown bread a pleasant sourness and aroma. Yeast white bread is baked in a random way - the yeast is placed together with flour and other components immediately in the main batch. Immediately before baking, the mixed population contained in the dough is stimulated to multiply by the addition of milk, water, sugar, flour. The resulting dough "fits", increasing in volume due to the intense release of CO 2 during the rapid reproduction of yeast fermenting carbohydrates.

Winemaking.A variety of microorganisms live on the surface and inside the berries, including a lot of yeast. Therefore, squeezed juice - wort - begins to ferment without additional yeast addition. Artisanal winemaking is based on this.
   The process of fermentation can be prevented primarily by acetic and lactic acid bacteria, unwanted yeast, and yeast-like fungi. To eliminate the risk of damage to wine material during the industrial production of wine, pre-grown and activated wine yeast is introduced into the grape must. The yeast races used, most often related to saccharomycetes, and the course of the fermentation process determine the type of wine. So, for example, in the manufacture of sherry special sherry yeast is used and barrels of wine are not filled to the brim (which is unacceptable in the manufacture of other wines).
The processes used in winemaking, studied in detail by Louis Pasteur. Yeast ferments the sugars found in grape juice (see chart above). Fermentation continues until the yeast has consumed all the sugar. Yeast forms alcohol only in the absence of oxygen or in the absence of oxygen. If there is a lot of oxygen, the yeast oxidizes the sugar completely to carbon dioxide and water. While fermentation proceeds violently, the emitted carbon dioxide protects the surface of the wort from interacting with atmospheric oxygen. When fermentation stops, the barrel of young wine must be sealed. If this is not done, acetic acid bacteria, using oxygen, will turn alcohol into acetic acid. This is how wine (or apple) vinegar is obtained. Based on the results of his research, Pasteur recommended that French winemakers observe microbiological purity in the preparation of wine: thoroughly wash barrels and fumigate wine with sulfur dioxide.

Brewing.Brewing, as well as distilling, is a traditional production in many countries of the world. As a rule, it is more industrialized than winemaking, and the yeast component is even more important here. The strains used are special types of saccharomycetes. Fermenting barley wort, yeast cells in a short time bring the alcohol content to 3-5%. In order to slow down the too intensive reproduction of yeast and to accumulate products that give the beer its taste (aldehydes, ketones, polyols), fermentation is carried out at low temperatures - 2–8 ° С. Under these conditions, further oxidation of aldehydes and alcohols almost does not occur.
   Many breweries are still equipped with open fermentation tanks, and only large factories have pressurized containers. Large yeast cells in the finished beer die and settle, a small proportion of them remain in suspension, and the continued fermentation of beer in storage tanks causes it to be saturated with carbon dioxide.

2. Penicillas

(student message)

Genus Penicillium ( Penicillium) refers to the order of hyphomycetes ( Hyphomycetales) from the class of imperfect mushrooms ( Deuteromycota) The natural habitat of these fungi is the soil, they are often found on a variety of substrates, mainly of plant origin.
Back in the XV – XVI centuries. in folk medicine, green mold was used to treat purulent wounds. In 1928, the English microbiologist Alexander Fleming noted that penicillium, accidentally entering the culture of staphylococcus, completely suppressed the growth of bacteria. These observations of Fleming formed the basis of the doctrine of antibiosis (antagonism between individual types of microorganisms). In the development of research on microbial antagonism, L. Pasteur, I.I. Mechnikov.
   The antimicrobial effect of green mold is due to a special substance - penicillin, released by this fungus into the environment. In 1940, penicillin was obtained in pure form by English researchers G. Flory and E. Cheyne, and in 1942, independently of them, by Soviet scientists Z. V. Ermolieva and T.I. Balezina. During World War II, penicillin saved the lives of hundreds of thousands of wounded. The demand for penicillin was so great that its production increased from several million units in 1942 to 700 billion units in 1945.
   Penicillin is used for pneumonia, sepsis, pustular skin diseases, tonsillitis, scarlet fever, diphtheria, rheumatism, syphilis, gonorrhea and other diseases caused by gram-positive bacteria.
   The discovery of penicillin laid the foundation for the search for new antibiotics and sources of their production. With the discovery of antibiotics, it became possible to successfully treat almost all infectious diseases caused by microbes.
   But green molds are successfully used not only in medicine. Of great importance are penicillas of the species P.roqueforti. In nature, they live in the soil. We are well acquainted with them by the group of cheeses characterized by marbling: Roquefort, whose homeland is France, Gorgonzola cheese from Northern Italy, Stilon cheese from England and others. All these cheeses have a loose structure, a specific “moldy” »Appearance (veins and spots of bluish-green color) and characteristic aroma. P.roqueforti   Needs a small amount of oxygen, tolerates high concentrations of carbon dioxide.
   In the preparation of soft French cheeses "Camembert", "Brie" and some others are used P.camamberti   and P.caseicolumwhich form a characteristic white “felt” coating on the surface of the cheese. under the influence of enzymes of these mushrooms, the cheese acquires juiciness, oiliness, specific taste and aroma.

3. Aspergillus

(student message)

(student message)

A huge amount of household garbage and agricultural and forestry waste is generated annually in the world. Wood and straw, as well as paper waste, which makes up almost half of the garbage, consists of three main components:

One should get rid of waste, on the one hand, as little as possible polluting the environment, and on the other hand, extracting as much energy and carbon from organic compounds from them as possible. At present, the waste is most often burned or disposed of untreated, without even getting thermal energy in the latter case.

However, alternative approaches based on the use of fungi in combination with other microorganisms are possible. One of the ways of recycling is the cultivation of edible mushrooms and fodder yeast on wood waste, but in total no more than 2% of organic waste is processed in this way.

For the decomposition of cellulose and lignin, it is preferable to use mushrooms, because the activity of the enzymes contained in them - cellulases and ligninases - is higher than that of bacterial enzymes, especially in an acidic environment that is characteristic of wood waste (bacteria prefer a slightly alkaline environment).

Work with tab. 1.

Table 1. Recycling Methods Using Mushrooms

6. Conclusions on the topic

There are no “good” and “bad” mushrooms; all of them are an integral part of the microworld, which ensures the circulation of substances in the biosphere.

Table 2. Properties of mushrooms

A person should study mushrooms in order to reduce or prevent the harm they cause and to usefully use their farming practices.

Securing the studied material

To consolidate the studied material, students perform two versions of test tasks.

Test tasks

Option 1

Write down the numbers of offers, mark the correct ones with a “+” sign.

Write down the numbers of the questions and write down the letters of the correct answers.

1. The science that studies the possibilities of using living organisms and biological processes in production is called:
   a) mycology; b) biotechnology; c) microbiology.

3. The combined group of unicellular mushrooms is called:
   a) bacteria; b) archebacteria; c) yeast; d) mucus.

4. Carnivorous fungi include:
   a) penicillus; b) aspergillus; c) yeast; d) arthrobotris.

5. Mushrooms play an important role in the recycling of garbage, household waste and agricultural waste:
   a) yes; b) no.

6. The first antibiotic obtained with the help of molds was:
   a) penicillin; b) tetracycline; c) levomycetin; g) streptomycin.

7. Microscopic mushrooms are used in the production of:
   a) enzymes; b) antibiotics; c) organic acids; d) all answers are correct.

1st option. 1+, 2–, 3–, 4–, 5+, 6+, 7–, 8+, 9–, 10+.

2nd option. 1b, 2g, 3c, 4g, 5a, 6a, 7g.

Homework

Examine the notes, prepare for the standings on the topic "Biotechnology".

As about objectsbiotechnologies can be performed: cells of microorganisms, animals and plants, transgenic animals and plants, as well as multicomponent enzyme systems of cells and individual enzymes.

The basis of most modern biotechnological industries is microbial synthesis, i.e., the synthesis of various biologically active substances with the help of microorganisms. Regardless of the nature of the facility, the primary step in the development of any biotechnological process is to obtain pure crops   organisms (if they are microbes), cells or tissues (if they are more complex organisms - plants or animals). Many stages of further manipulations with the latter (i.e., with the cells of plants or animals) are the principles and methods used in microbiological industries. From the methodological point of view, both microbial cell cultures and tissue cultures of plants and animals practically do not differ from microbial cultures. World m organisms   extremely diverse. In n. time is known for more than 100 thousand of their various types. it prokaryotes(bacteria, actinomycetes, rickettsia, cyanobacteria) and part e ukriot   (yeast, filamentous mushrooms, some protozoa and algae). With a wide variety of microorganisms, an important problem is the correct choice of the organism that is able to provide the desired product, i.e., serve industrial purposes. Microorganisms:

1) Industrial : E. coli ( E. coli), hay stick ( You. sub-tilis) and baker's yeast ( S. cerevisiae) Usually they are super producers. To obtain super-producers, they carry out genetic selection work, genetic engineering approaches (introducing human genes. Into the bacterium: interferon, insulin genes, etc.). PS should be patented.

2) Basic   used in limited numbers, classified as GRAS   ("generally recognized as safe" is generally considered safe) - bacteria Bacillus subtilis, Bacillus amylolique-faciens,   other types of bacilli and lactobacilli, species Streptomycesmushrooms Aspergillus, Penicillium, Mucor, Rhizopusyeast Saccharomyces and   dr . GRAS- microorganisms are non-pathogenic, non-toxic and generally do not form antibiotics, therefore, when developing a new biotechnological process, one should focus on these microorganisms.

3) Model-   bacilli (producers of proteolytic enzymes). There are catalogs of model micr.

Main criterion   when choosing a biotechnological object is the ability to synthesize the target product. microorganisms must (requirements):

Have a high growth rate;

Dispose of cheap substrates necessary for their life;

Be resistant to extraneous microflora, i.e. possess high competitiveness. (requirements): the ability to grow on cheap substrates, high economic coefficient, minimal education of by-products (toxic metabolites, allergens)

All of the above provides a significant reduction in the cost of manufacturing the target product. The following are examples aimed at illustrating the foregoing.

1. Unicellular organisms   characterized by higher growth rates and synthetic processes,

2. Particular attention as objects of biotechnological developments are photosynthetic microorganismsusing the energy of sunlight in their life.

3. thermophilic microorganismsgrowing at 60-80 ° C. This property of them is an almost insurmountable obstacle to the development of extraneous microflora.

24. Advantages of microorganisms over other objects in solving modern biotechnological problems:

· Small size

Ubiquitous

· Various types of metabolism

· Phototrophs

· Occupy a small volume (in 1 ml up to 1 billion individuals)

· High fission rate, fast growth

· Able to live in various conditions.

Photosynthetic organisms are promising as producers of ammonia, hydrogen, and protein.

Thermophilic microorganisms growing at 60-80 degrees. This is a reliable protection against contamination. Enzymes synthesized by thermophiles, charac. increased resistance to heat, but at the same time they are inactive at ordinary temperatures.

Biotechnology as a science and industry. The subject, goals and objectives of biotechnology, communication with fundamental disciplines.

Biotechnology is a technological process using biotechnological systems - living organisms and components of a living cell. Systems can vary from microbes and bacteria to enzymes and genes. Biotechnology is a production based on the achievements of modern science: genetic engineering, physical chemistry of enzymes, molecular diagnostics and molecular biology, selection genetics, microbiology, biochemistry, chemistry of antibiotics.

In the field of pharmaceutical production, biotechnology supplants traditional technologies and opens up fundamentally new opportunities. Biotechnological methods produce genetically engineered proteins (interferons, interleukins, insulin, hepatitis vaccines, etc.), enzymes, diagnostic tools (drug test systems, drugs, hormones, etc.), vitamins, antibiotics, biodegradable plastics, biocompatible materials.

Immune biotechnology, with the help of which single cells are recognized and isolated from mixtures, can be used not only directly in medicine for diagnosis and treatment, but also in scientific research, in the pharmacological, food and other industries, as well as used to obtain preparations synthesized by cells body defense system.

Currently, the achievements of biotechnology are promising in the following sectors:

in industry (food, pharmaceutical, chemical, oil and gas) - the use of biosynthesis and biotransformation of new substances based on genetically engineered strains of bacteria and yeast with desired properties based on microbiological synthesis;

in ecology - improving the efficiency of environmental protection of plants, developing environmentally friendly wastewater treatment technologies, utilizing agricultural wastes, designing ecosystems;

in the energy sector - the use of new sources of bioenergy obtained on the basis of microbiological synthesis and simulated photosynthetic processes, bioconversion of biomass into biogas;

in agriculture - the development of transgenic agricultural crops, biological plant protection products, bacterial fertilizers, microbiological methods, soil remediation in the field of crop production; in the field of animal husbandry - the creation of effective feed preparations from plant, microbial biomass and agricultural waste, animal reproduction based on embryogenetic methods;

In medicine - the development of medical biologics, monoclonal antibodies, diagnostics, vaccines, the development of immunobiotechnology in the direction of increasing the sensitivity and specificity of the immunoassay of diseases of an infectious and non-infectious nature.

Compared to chemical technology, biotechnology has the following main advantages:

The ability to obtain specific and unique natural substances, some of which (for example, proteins, DNA) have not yet been obtained by chemical synthesis;

Conducting biotechnological processes at relatively low temperatures and pressures;

Microorganisms have significantly higher growth and accumulation rates of cell mass than other organisms. For example, using microorganisms in a fermenter with a volume of 300 m 3 per day, 1 ton of protein can be produced (365 t / year). In order to develop the same amount of protein per year with the help of cattle, you need to have a herd of 30,000 heads. If you use legumes, such as peas, to obtain such a rate of protein production, you will need to have a pea field of 5400 ha;

As raw materials in biotechnology processes, cheap waste from agriculture and industry can be used;

Biotechnological processes in comparison with chemical ones are usually more environmentally friendly, have less harmful wastes, are close to natural processes occurring in nature;

As a rule, technology and equipment in biotechnological industries are simpler and cheaper.

The priority task for biotechnology is the creation and development of the production of medicines for medicine: interferons, insulins, hormones, antibiotics, vaccines, monoclonal antibodies and others, allowing early diagnosis and treatment of cardiovascular, malignant, hereditary, infectious, including viral diseases.

The concept of "biotechnology" is collective and covers areas such as fermentation technology, the use of biofactors using immobilized microorganisms or enzymes, genetic engineering, immune and protein technologies, technology using cell cultures of both animal and plant origin.

Biotechnology is a combination of technological methods, including genetic engineering, using living organisms and biological processes for the production of medicines, or the science of the development and application of living systems, as well as non-living systems of biological origin in the framework of technological processes and industrial production.

Modern biotechnology is chemistry, where the change and transformation of substances occurs through biological processes. In intense competition, two chemistry are successfully developing: synthetic and biological.

Bioobjects as a means of production of therapeutic, rehabilitation, prophylactic and diagnostic tools. Classification and general characteristics of bioobjects.

The objects of biotechnology are viruses, bacteria, fungi - micromycetes and macromycetes, protozoal organisms, cells (tissues) of plants, animals and humans, some biogenic and functionally similar substances (for example, enzymes, prostaglandins, pectins, nucleic acids, etc.). Therefore, biotechnology objects can be represented by organized particles (viruses), cells (tissues) or their metabolites (primary, secondary). Even when using a biomolecule as an object of biotechnology, its initial biosynthesis is carried out in most cases by the corresponding cells. In this regard, we can say that the objects of biotechnology belong either to microbes, or to plant and animal organisms. In turn, the body can be figuratively characterized as a system of economical, complex, compact, self-regulating and, therefore, targeted biochemical production, which proceeds steadily and actively while optimally maintaining all the necessary parameters. From this definition it follows that viruses are not organisms, but according to the content of the molecules of heredity, adaptability, variability and some other properties, they belong to representatives of wildlife.

As can be seen from the diagram, biotechnology objects are extremely diverse, their range extends from organized particles (viruses) to humans.

Currently, most of the biotechnological objects are microbes belonging to the three kingdoms (non-nuclear, pre-nuclear, nuclear) and five kingdoms (viruses, bacteria, fungi, plants and animals). Moreover, the first two kingdoms consist exclusively of microbes.

Microbes among plants are microscopic algae (Algae), and among animals are microscopic algae (Protozoa). From eukaryotes, fungi belong to microbes and, with certain reservations, lichens, which are natural symbiotic associations of microscopic fungi and microalgae or fungi and cyanobacteria.

Acaryota - non-nuclear, Rosaruota - pre-nuclear and EU-uota - nuclear (from Greek a - no, rro - to, eu - good, full, saun - the core). Organized particles belong to the first - viruses and viroids, to the second - bacteria, to the third - all other organisms (fungi, algae, plants, animals).

Microorganisms form a huge number of secondary metabolites, many of which have also found application, for example, antibiotics and other mammalian cell homeostasis correctors.

Probiotics - preparations based on the biomass of certain types of microorganisms are used for dysbiosis to normalize the microflora of the gastrointestinal tract. Microorganisms are also needed in the production of vaccines. Finally, microbial cells can be transformed by genetic engineering into producers of species-specific protein hormones for humans, protein factors of nonspecific immunity, etc.

Higher plants are traditional and still the most extensive source of obtaining medicines. When using plants as biological objects, the main attention is focused on the issues of cultivating plant tissues on artificial media (callus and suspension cultures) and opening up new perspectives.

Macrobioobjects of animal origin. Man as a donor and object of immunization. Mammals, birds, reptiles, etc.

In recent years, in connection with the development of recombinant DNA technology, the importance of such a biological object as a person is rapidly growing, although at first glance this seems paradoxical.

However, a person became a bioobject from the standpoint of biotechnology (using bioreactors) only after realizing the possibility of cloning his DNA (more precisely, its exons) in the cells of microorganisms. Due to this approach, the shortage of raw materials for obtaining species-specific human proteins was eliminated.

Important in biotechnology are macro objectswhich include various animals and birds. In the case of the production of immune plasma, a person acts, in addition, as an object of immunization.

To obtain various vaccines, organs and tissues, including embryonic ones, of various animals and birds are used as objects for the reproduction of viruses: It should be noted that the term "donor"in this case, the bio-object that supplies the material for the production process of the medicinal product without damage to its own life is indicated, and the term Donor- a bio-object in which the intake of material for the production of a medicinal product is incompatible with the continuation of life.

Of the embryonic tissues, chicken embryonic tissues are the most widely used. Of particular benefit are chicken embryos (when available) of ten to twelve days of age, used primarily for the reproduction of viruses and the subsequent manufacture of viral vaccines. Chicken embryos were introduced into virological practice in 1931 by G. M. Woodruff and E. W. Goodpascher. Such embryos are also recommended for identifying, identifying and determining the infectious dose of viruses, for obtaining antigenic preparations used in serological reactions.

Incubated at 38 ° C, chicken eggs are ovoscopic (transparent), discarded, “transparent” unfertilized specimens and kept fertilized, in which the filled blood vessels of the chorionallantoic membrane and the movement of the embryos are clearly visible.

Infection of embryos can be done manually and automatically. The latter method is used in large-scale production of, for example, influenza vaccines. Material containing viruses is injected with a syringe (syringe battery) into various parts of the embryo (embryos).

All stages of work with chicken embryos after ovoscopy are carried out under aseptic conditions. The material for infection may be a suspension of ground brain tissue (for rabies virus), liver, spleen, kidneys (for chlamydia ornithosis), etc. In order to decontaminate the viral material from bacteria or to prevent bacterial contamination, appropriate antibiotics can be used for example, penicillin with any aminoglycoside of the order of 150 IU each per 1 ml of a suspension of virus-containing material. To combat fungal infection of embryos, it is advisable to use some antibiotics-polyenes (nystatin, amphotericin B) or certain benzimidazole derivatives (for example, dactarin, etc.).

Most often, a suspension of viral material is injected into the allantoic cavity or, less commonly, on the chorionallantoic membrane in an amount of 0.05-0.1 ml, piercing a disinfected shell (e.g., iodinated ethanol) to the calculated depth. After that, the hole is closed with molten paraffin and the embryos are placed in a thermostat, which maintains the optimum temperature for the reproduction of the virus, for example 36-37.5 ° C. The duration of incubation depends on the type and activity of the virus. Usually, after 2-4 days, you can observe a change in the membranes with subsequent death of the embryos. Infected embryos are monitored daily 1-2 times (ovoscopy, turn the other side). Dead embryos are then transferred to the viral material collection unit. There they are disinfected, the allantoic fluid with the virus is aspirated and transferred to sterile containers. Inactivation of viruses at a certain temperature is usually carried out using formalin, phenol or other substances. Using high-speed centrifugation or affinity chromatography (see), it is possible to obtain highly purified viral particles.

The collected viral material, passed the appropriate control, is subjected to freeze drying. The following indicators are subject to control: sterility, safety and specific activity. With regard to sterility, they mean the absence of: a living homologous virus in the killed vaccine, bacteria and fungi. Harmlessness and specific activity are evaluated in animals and only after that the vaccine is allowed to be tested on volunteers or volunteers; after successful clinical testing, the vaccine is allowed to be used in wide medical practice.

For chicken embryos, for example, livinginfluenza vaccine. It is intended for intranasal administration (persons over 16 years old and children from 3 to 15 years old). A vaccine is a dried allantoic fluid taken from virus-infected chicken embryos. The type of virus is selected according to the epidemiological situation and forecasts. Therefore, drugs can be produced in the form of a single vaccine or a divacine (for example, including viruses A2 and B) in ampoules with 20 and 8 vaccination doses for the respective population groups. The dried mass in ampoules usually has a light yellow color, which persists after dissolving the contents of the ampoule in boiled cooled water.

Live influenza vaccines for adults and children are also prepared for oral administration. Such vaccines are special vaccine strains, the reproduction of which occurred within 5-15 passages (no less and no more) on the kidney tissue culture of chicken embryos. They are released dry in vials. When dissolved in water, the color changes from light yellow to reddish.

Of the other viral vaccines obtained on chicken embryos, it is possible to call anti-parotid, against yellow fever.

Of the other embryonic tissues, the embryos of mice or other mammalian animals are used, as well as aborted human fetuses.

Embryonic transplantable tissues are accessible after trypsin treatment, since a large amount of intercellular substances (including non-proteinaceous ones) is not yet formed in such tissues. Cells are separated and, after necessary treatments, they are cultured in special media in a monolayer or in a suspended state.

Tissues that are isolated from animals after birth are classified as mature.The greater their age, the more difficult they are to be cultivated. However, after successful cultivation, they then “align” and differ little from embryonic cells.

In addition to polio, specific prophylaxis with live vaccines is carried out with measles. Measles live dry vaccinemade from a vaccine strain, the reproduction of which was carried out on cell cultures of Guinea pig kidneys or Japanese quail fibroblasts.

Bioobjects of plant origin. Wild plants and plant cell cultures.

Plants are characterized by: the ability to photosynthesis, the presence of cellulose, starch biosynthesis.

Algae are an important source of various polysaccharides and other biological active substances. Oii reproduce vegetatively, asexually and sexually. As biological objects are not used enough, although, for example, kelp called sea kale is produced by industry in various countries. Agar agar and alginates derived from algae are well known.

Cells of the highest rasteyi. Higher plants (about 300,000 species) are differentiated multicellular, mainly terrestrial organisms. Of all the tissues, only meristematic tissues are capable of division, and due to them all other tissues are formed. This is important for obtaining cells, which then must be included in the biotechnological process.

Meristem cells that linger at the embryonic stage of development throughout the life of the plant are called initial, others gradually differentiate and turn into cells of various permanent tissues - the final cells.

Depending on the topology in the plant, meristems are divided into apical, or apical (otl. Walnut - top), lateral, or lateral (from Lat. Lateralis - lateral) and intermediate, or intercalar (from Lat. Intercalaris - interstitial, inserted.

Typeotency   - this is the property of somatic plant cells to fully realize their development potential up to the formation of a whole plant.

Any type of plant can produce, under appropriate conditions, an unorganized mass of dividing cells - callus (ref. Callus - corn), especially with the inducing effect of plant hormones. Mass production of calluses with further regeneration of shoots is suitable for large-scale plant production. In general, callus is the main type of plant cell cultured on a nutrient medium. Callus tissue from any decay can be recultivated for a long time. In this case, the initial plants (including meristematic ones), differentiate and despecialize, but are induced to divide, forming a primary callus.

In addition to growing calluses, it is possible to cultivate the cells of some plants in suspension cultures. Protoplasts of plant cells also appear to be important bioobjects. The methods for their preparation are fundamentally similar to the methods for producing bacterial and fungal protoplases. Subsequent cell-based experiments with them are attractive for possible valuable results.

Bioobjects are microorganisms. The main groups of biologically active substances.

The objects of biotechnology are viruses, bacteria, fungi - micromycetes and macromycetes, protozoal organisms, cells (tissues) of plants, animals and humans, some biogenic and functionally similar substances (for example, enzymes, prostaglandins, lectins, nucleic acids, etc.). Therefore, biotechnology objects can be represented by organized particles (viruses), cells (tissues) or their metabolites (primary, secondary). Even when using a biomolecule as an object of biotechnology, its initial biosynthesis is carried out in most cases by the corresponding cells. In this regard, we can say that the objects of biotechnology belong either to microbes, or to plant and animal organisms. In turn, the body can be figuratively characterized as a system of economical, complex, compact, self-regulating and, therefore, purposeful biochemical production, which proceeds stably and actively while optimally maintaining all the necessary parameters. From this definition it follows that viruses are not organisms, but according to the content of the molecules of heredity, adaptability, variability and some other properties, they belong to representatives of wildlife.

Currently, most of the biotechnological objects are microbes belonging to the three kingdoms (non-nuclear, pre-nuclear, nuclear) and five kingdoms (viruses, bacteria, fungi, plants and animals). Moreover, the first two kingdoms consist solely of microbes.

The cells of fungi, algae, plants and animals have a real nucleus that is delimited from the cytoplasm and therefore they are referred to as eukaryotes.

Bioobjects are macromolecules with enzymatic activity. Use in biotechnological processes.

Recently, a group of enzyme preparations has received a new direction of application - it is engineering enzymology, which is a branch of biotechnology where the enzyme acts as a bioobject.

Organotherapy, i.e. treatment with organs and preparations from animal organs, tissues and excreta, for a long time rested on deep empiricism and conflicting ideas, occupying a prominent place in medicine of all times and peoples. Only in the second half of the XIX century, as a result of the successes achieved by biological and organic chemistry, and the development of experimental physiology, did organotherapy become a scientific basis. This is due to the name of the French physiologist Brown-Secard. Particular attention was paid to the work of Brown-Secar related to the introduction into the human body of extracts from the testes of the bull, which had a positive effect on performance and well-being.

The first official drugs (GF VII) were adrenaline, insulin, pituitrin, pepsin and pancreatin. Subsequently, as a result of extensive research conducted by Soviet endocrinologists and pharmacologists, it became possible to consistently expand the range of official and unofficial organ preparations.

However, some amino acids are obtained by chemical synthesis, for example glycine, as well as D-, L-methionine, the D-isomer of which is low toxic, therefore a methionine-based medicine contains D- and L-forms, although the drug is used abroad in medicine containing only the L-form of methionine. There, the racemic mixture of methionine is separated by bioconversion of the D-form into the L-form under the influence of special enzymes of living cells of microorganisms.

Immobilized enzyme preparations have a number of significant advantages when used for practical purposes compared with native predecessors. First, a heterogeneous catalyst can be easily separated from the reaction medium, which makes it possible: a) to stop the reaction at the right time; b) reuse the catalyst; c) receive a product that is not contaminated with the enzyme. The latter is especially important in a number of food and pharmaceutical industries.

Secondly, the use of heterogeneous catalysts allows the enzymatic process to be carried out continuously, for example in flow columns, and to control the rate of the catalyzed reaction, as well as the yield of the product by changing the flow rate.

Thirdly, immobilization or modification of the enzyme contributes to a targeted change in the properties of the catalyst, including its specificity (especially in relation to macromolecular substrates), the dependence of catalytic activity on pH, ionic composition and other environmental parameters and, very importantly, its stability with respect to to various denaturing effects. Note that a major contribution to the development of general principles for the stabilization of enzymes was made by Soviet researchers.

Fourth, the immobilization of enzymes makes it possible to regulate their catalytic activity by changing the properties of the carrier under the influence of certain physical factors, such as light or sound. On this basis, mechano- and sound-sensitive sensors, amplifiers of weak signals and silver-free photographic processes are created.

As a result of the introduction of a new class of bioorganic catalysts - immobilized enzymes, new, previously inaccessible ways of development have opened before applied enzymology. Just listing the areas in which immobilized enzymes are used could take up a lot of space.

Directions for improving bioobjects by selection and mutagenesis methods. Mutagens. Classification. Characteristic. The mechanism of their action.

That mutations are the primary source of organisms' variability, creating the basis for evolution. However, in the second half of the XIX century. another source of variability was discovered for microorganisms - the transfer of foreign genes - a kind of "genetic engineering of nature."

For a long time, the concept of mutation was referred only to chromosomes in prokaryotes and chromosomes (nucleus) in eukaryotes. Currently, in addition to chromosomal mutations, the concept of cytoplasmic mutations has also appeared (plasmid mutations in prokaryotes, mitochondrial and plasmid mutations in eukaryotes).

Mutations can be caused both by rearrangement of the replicon (a change in the number and order of genes in it), as well as changes within the individual gene.

In relation to any biological objects, but especially often in the case of microorganisms, the so-called spontaneous mutations are detected, which are found in a population of cells without special effects on it.

According to the severity of almost any trait, cells in the microbial population make up the variation series. Most cells have a moderate severity of the trait. Deviations of “+” and “-” from the average value are found in the population more rarely, the larger the deviation in either direction (Fig. I). The initial, simplest approach to improving the bioobject was to select deviations “+” (assuming that these deviations correspond to the interests of production). In the new clone (genetically homogeneous offspring of one cell; on a solid medium — a colony) obtained from a cell with a deviation of “+”, selection was again carried out according to the same principle. However, such a procedure, when it is repeated many times, quickly loses its effectiveness, that is, the deviations “+” in the new clones become smaller and smaller.