1 Introduction 3 2 Experimental part 4 2.1 The concept of a bioobject 4 2.2 Improvement of bioobjects by mutagenesis and selection methods 7 2.3 Genetic engineering methods 12 3 Conclusions and suggestions 24 References 25

Introduction

The tasks of modern breeding include the creation of new and improvement of existing plant varieties, animal breeds and strains of microorganisms. The theoretical basis of breeding is genetics, since it is the knowledge of the laws of genetics that makes it possible to purposefully control the appearance of mutations, predict the results of crossing, and correctly select hybrids. As a result of the application of knowledge in genetics, it was possible to create more than 10,000 varieties of wheat based on several initial wild varieties, to obtain new strains of microorganisms that secrete food proteins, medicinal substances, vitamins, etc. In connection with the development of genetics, selection received a new impetus to development. Genetic engineering allows organisms to be purposefully modified. Genetic engineering is used to obtain the desired qualities of a modified or genetically modified organism. Unlike traditional selection, during which the genotype is only indirectly changed, genetic engineering allows you to directly interfere with the genetic apparatus, using the technique of molecular cloning. Examples of the application of genetic engineering are the production of new genetically modified varieties of crops, the production of human insulin by using genetically modified bacteria, the production of erythropoietin in cell culture, etc.

Conclusion

Genetic engineering is a promising area of ​​modern genetics, which is of great scientific and practical importance and underlies modern biotechnology. To obtain the necessary target product of genetic engineering, as well as for economic benefits, it is necessary to use methods such as mutagenesis and selection. These methods are widely used in the production of many medicinal substances (for example, the production of human insulin through the use of genetically modified bacteria, the production of erythropoietin in cell culture, etc.), the production of new genetically modified varieties of crops, and much more. The application of the laws of genetics allows you to correctly manage the methods of selection and mutation, predict the results of crossing, and correctly select hybrids. As a result of applying this knowledge, it was possible to create more than 10,000 varieties of wheat based on several original wild varieties, to obtain new strains of microorganisms that secrete food proteins, medicinal substances, vitamins, etc.

Bibliography

1. Blinov V. A. General biotechnology: a course of lectures. Part 1. FGOU VPO "Saratov State Agrarian University". Saratov, 2003. - 162 p. 2. Orekhov S.N., Katlinskii A.V. Biotechnology. Proc. allowance. - M.: Publishing Center "Academy", 2006. - 359 p. 3. Katlinsky A.V. Course of lectures on biotechnology. – M.: Publishing house MMA im. Sechenov, 2005. - 152 p. 4. Bozhkov A. I. Biotechnology. Fundamental and industrial aspects. - H.: Fedorko, 2008. - 363 p. 5. Popov V.N., Mashkina O.S. Principles and basic methods of genetic engineering. Proc. allowance. Publishing and Printing Center of VSU, 2009. - 39 p. 6. Shchelkunov S.N. genetic engineering. Study guide allowance. - Novosibirsk: Sib. univ. publishing house, 2004. - 496 p. 7. Glick B. Molecular biotechnology: principles and applications /B. Glick, J. Pasternak. - M. : Mir, 2002. - 589 p. 8. Zhimulev I.F. General and molecular genetics / I.F. Zhimulev. - Novosibirsk: Publishing house Novosib. un-ta, 2002. - 458 p. 9. Rybchin V.N. Fundamentals of genetic engineering / V.N. Rybchin. - St. Petersburg: Publishing House of St. Petersburg State Technical University, 1999. - 521p. 10. Electron. textbook allowance / N. A. Voynov, T. G. Volova, N. V. Zobova and others; under scientific ed. T. G. Volovoy. - Krasnoyarsk: IPK SFU, 2009.

Bioobject- this is a producer that biosynthesizes the desired product, or a catalyst, an enzyme that catalyzes its inherent reaction.

Requirements for biological objects

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

It should be borne in mind that when favorable conditions are created for a selected biological object of biotechnology, the same conditions may turn out to be favorable, for example, for microbes - contaminants or pollutants. Representatives of the contaminating microflora are viruses, bacteria and fungi found in cultures of plant or animal cells. In these cases, microbes-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 destruction by banal saprophytic (not pathogenic) microflora, which can penetrate into 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 biological object, in practice, either natural organized particles (phages, viruses) and cells with natural genetic information, or cells with artificially given genetic information are used, that is, in any case, cells are used, be it a microorganism, a plant, animal or person. For example, we can name the process of obtaining the polio virus on a culture of monkey kidney cells 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 to be used in an immobilized state. The source of enzymes is also isolated cells or their specialized associations in the form of tissues, from which the necessary biocatalysts are isolated.

Classification of biological objects

1) Macromolecules

Enzymes of all classes (often hydrolases and transferases); including in an immobilized form (associated with the carrier) providing multiple use and standardization of repetitive production cycles;

DNA and RNA - in an isolated form, as part of foreign cells.

2) Microorganisms

Viruses (with attenuated pathogenicity are used to produce vaccines);

Prokaryotic and eukaryotic cells are producers of primary metabolites: amino acids, nitrogenous bases, coenzymes, mono- and disaccharides, enzymes for replacement therapy, etc.); -producers of secondary metabolites: antibiotics, alkaloids, steroid hormones, etc.;

Normoflora - the biomass of certain types of microorganisms used for the prevention and treatment of dysbacteriosis;

Pathogens of infectious diseases - sources of antigens for the production of vaccines;

Transgenic m / o or cells - producers of species-specific protein hormones for humans, protein factors of nonspecific immunity, etc.

3) Macroorganisms

Higher plants are raw materials for obtaining biologically active substances;

Animals - mammals, birds, reptiles, amphibians, arthropods, fish, molluscs, humans;

transgenic organisms.

As biological objects or systems that biotechnology uses, first of all, it is necessary to name unicellular microorganisms, as well as animal and plant cells. The choice 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 course of life: proteins, fats, carbohydrates, vitamins, nucleic acids, amino acids, antibiotics, hormones, antibodies, antigens, enzymes, alcohols, etc. Many of these products, are extremely necessary in human life, are not yet available for obtaining by "non-biotechnical" methods due to the scarcity or high cost of raw materials or the complexity of technological processes.

2. Cells reproduce extremely quickly. Thus, a bacterial cell divides every 20–60 minutes, a yeast cell divides every 1.5–2 hours, and an animal cell divides every 24 hours, which makes it possible to artificially grow huge amounts of biomass on a relatively cheap and non-deficient nutrient media on an industrial scale in a relatively short time. microbial, animal or plant cells. For example, in a bioreactor with a capacity of 100 m 3 for 2-3 days. 10 16 -10 18 microbial cells can be grown. During the life of cells during their cultivation, a large number of valuable products enter the environment, and the cells themselves are pantries of these products.

3. Biosynthesis of complex substances such as proteins, antibiotics, antigens, antibodies, etc. is much more economical and technologically more accessible than chemical synthesis. At the same time, the feedstock for biosynthesis, as a rule, is simpler and more accessible than the feedstock for other types of synthesis. For biosynthesis, waste from the agricultural, fishing, food industry, vegetable raw materials, yeast, wood, molasses, etc.) is used.

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 technology, etc.

Anthropogenic impact on the biosphere is integral to the development of civilization. Plowing of land, deforestation, "trampling" of the steppes constantly accompany the history of mankind. It is appropriate to recall the destruction of certain species of animals and plants and the resettlement of some species from their native habitats.

In connection with the particular relevance of the problem of the influence of industry on the biosphere, let us consider what biotechnological production looks like in this regard. First of all, it is science-intensive and, in comparison with chemical-technological production, is more efficient, since the cell of the producer (bio-object) is a “balanced complex of biocatalysts” that works more productively than systems of sequential chemical reactions with inorganic catalysts.

The consumption of energy resources and water by the biotechnology industry is a fraction of a percent of that consumed by the modern chemical industry. Emission of gaseous wastes of biotechnological enterprises into the atmosphere does not exceed even a tenth of a percent of the emissions from the industry as a whole. It is biotechnological production that is most acceptable in modern conditions, however, it also has specific environmental problems and, accordingly, is being improved in the following directions:

Creation and use of more active biological objects-producers (as a result, there will be less waste per unit of production!);

Replacement of media and reagents with less scarce ones;

Immobilization of biological objects (both cells and enzymes), their repeated use to reduce waste;

Implementation of membrane technology at the stage of isolation and purification of the target product (reducing the amount of organic solvents used to avoid aggressive conditions at some stages of the production process);

Compliance with GMP rules.

Let us briefly consider the problems related to the elimination (utilization) or treatment of industrial waste of a traditional biotechnological enterprise.

solid waste. First of all, these include the mycelium (biomass) of the producer after its separation from the culture fluid and the target product. The amount of mycelium that has to be dealt with can be visualized based on the fact that the volume of the discharge of an industrial fermenter is 50-100 m 3 of a thick, viscous (due to the presence of mycelium) liquid. Considering that the enterprise has a number of fermenters, and the fermentation cycle lasts about a week, it can be concluded that this type of solid waste at one (large) enterprise amounts to hundreds of tons per year. It should be taken into account that the mycelium also contains residual amounts of the target product, and these are, as a rule, biologically highly active substances.

Currently, solid waste is eliminated by processing mycelium. It is mixed with soil and placed in pits with concrete substrates. Each such hole is left closed

for several years. During this time, soil microorganisms are subjected to organic matter mycelium to enzymatic cleavage, using them to build "their" biomass. In fact, compost is formed, while the organic part of the mycelium decomposes. A concrete underlayment in these "compost pits" is necessary to prevent the undecomposed soluble organic matter of the mycelium from entering the groundwater and rainwater reservoirs. Usually, special areas are allocated for compost pits on the territory of the enterprise. It should be noted that the export of dried mycelium (its mass decreases by 10-100 times compared to the original) to city dumps is prohibited.

Attempts to use mycelium for various purposes have not yet been successful in general, but a low-waste technology has already been created in laboratory conditions. The total lipid fraction was extracted from the mycelium of the actinomycete of the tetracycline producer and used as a defoamer in the next production cycle in the production of tetracycline produced by the producer belonging to the same strain. In some cases (with limited pastures), the sterilized and milled biomass of some microorganisms is used as an additive in the feed of farm animals. Mycelium of fungi and actinomycetes (waste in the production of antibiotics) improves the quality of some building materials (claydite slabs, bricks, etc.), increasing their strength. But for economic reasons, it is impractical to produce these materials.

Liquid waste. AT In the case of biotechnological production, liquid wastes are effluents and waste liquid, mainly this is a cultural liquid after the mycelium is separated from it and the target product is extracted. The total annual volume of culture liquid, which must be purified, is tens of thousands of cubic meters for one enterprise. The degree of purification, controlled by various methods, must be such that the purified liquid can be discharged into open water bodies.

There are different cleaning schemes. In almost all of them, microorganisms play a key role (biological treatment). We present one of these schemes. The first component of the purification system is a reinforced concrete sump where the spent culture liquid enters. At the bottom of the sump, pipes are laid through which the sediment is sucked off. At this stage, approximately 40% of contaminants are removed from the culture liquid.

The next section of the purification system consists of one or more aerotanks located one after the other - tanks with pipes passing along the bottom, from which air comes out in the form of bubbles, passing through the entire thickness of the liquid, as a result it is saturated with oxygen. Air contributes to the intensive flow of oxidative processes. The key feature of the aerotank is the presence in it of the so-called "activated sludge" (artificial biocenosis - a community of microorganisms that oxidize organic substances dissolved in the liquid to CO 2 and H 2 O), which gradually forms during the operation of the enterprise.

The species composition of the activated sludge biocenosis at different enterprises may vary slightly, since the latter depends on the oxidized substrates. As a rule, it is dominated by representatives of the genus Pseudomonas (70%). This is followed by microorganisms united in the genus Bacterium (20%). The remaining 10% are representatives of the genera Bacillus, Sarcina and other microorganisms. When characterizing activated sludge as a biocenosis or as a supraorganismal interspecific community in relation to the treatment of wastewater from biotechnological production, three important circumstances should be noted.

First, strains of the genus Pseudomonas play a fundamental role here. However, this genus should not be reduced to Pseudomonas acruginosa, a known causative agent of dangerous wound infections. Under natural conditions, the genus Pseudomonas is represented by a large number of species that are not dangerous to humans. It is non-pathogenic strains that are part of activated sludge. These microorganisms are characterized by a wide range of oxidative enzymes. Preparations consisting of Pseudomonas cells are used in the elimination of pollution caused by oil spills. Figuratively speaking, exotic substrates, such as annular hydrocarbons, also undergo oxidation. In addition, the shell of saprophytic Pseudomonas species included in activated sludge has its own characteristics at the level of porin channels, which facilitate the access of substrates to oxidative enzymes.

Secondly, the transformation of some substrates into CO 2 and H 2 O is carried out due to the sequential action of the enzymes of various microorganisms on them. In other words, one enzyme system converts a particular compound into intermediates, while another catalyzes the further degradation of these intermediates. This emphasizes that activated sludge functions as a complex of microorganisms.

Thirdly, it should be borne in mind that the wastewater of some industries (in particular, enterprises of the antibiotic industry) may contain residual amounts of antimicrobial substances. This means that the microorganisms in the aerotanks are constantly in contact with them, i.e. conditions are created for the selection of resistant forms. But there are cases when the concentration of antimicrobial substances in the treated liquid waste may be unusually high and cause the death of activated sludge cells.

This requires control over the state of activated sludge. After a section with an aerotank or several successively located aerotanks and a secondary settling tank, the “post-treatment unit” is fundamentally important for the liquid waste system. In it, the cultural liquid, in which approximately 10% of the original content of organic substances remains (as a rule, these are difficult-to-oxidize substances), is passed through biofilters - films with immobilized cells of microorganisms with the highest oxidizing activity. Quite often, these cells belong to genetically engineered strains containing plasmids carrying genes for oxidative enzymes (destruction enzymes). Such purposefully obtained "destructor strains" are capable of oxidizing substances that are difficult to oxidize and destroy the remaining 10% of contaminants in the treated liquid.

The immobilization of cells of such strains in biofilms is rational in view of the fact that during the free reproduction of these cells, artificially increased oxidative activity can be lost due to back mutations or loss of plasmids. In this case, genetic engineering and engineering enzymology seem to be “combined” in the “post-treatment block”. A post-purified liquid that meets the official criteria for drinking water (one of the accepted methods of toxicity control in this case is the suppression of microscopic

crustacean Daphnia magna), is chlorinated and then enters open water bodies.

Regarding the operation of biological wastewater treatment systems in different modes, it should be noted that at maximum (“shock”) loads, various difficulties may arise. During such working periods, highly active destructor strains (“bacterial starters”) are introduced into the aerotanks, which makes it possible to significantly increase the throughput of the liquid waste treatment system. For this purpose, special preparations are recommended for biotechnological enterprises of various profiles: "Phenobac" - for the utilization of hydrocarbons, "Thermobac" - for the oxidation of polysaccharides, "Polibac" - for release from synthetic detergents, etc. The approximate dose of "bacterial starter" from living cells is about 100 mg per 1 m 3 of waste fluid.

In conclusion, we note the possible variety of schemes for the biological utilization of liquid waste. So, in addition to aerobic purification, the scheme can include: an anaerobic purification stage, stages using sorbents (activated carbon, zeolites, etc.), stages using electrochemical methods (for example, electrocoagulation).

Gaseous waste. Gas emissions are purified from organic compounds at temperatures from 300 to 1,000 °C in columns with inorganic catalysts. In this case, the volatile "organics" turns into CO 2 . In some cases, biological filters based on microorganisms that oxidize organic substances to CO 2 are used.

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Department of Microbiology and Biochemistry

Guidelines

To be completed control work

on the topic: "Application of genetic engineering methods in biotechnology"

By discipline: Introduction to Biotechnology

for direction 020200.62 "Biology

Form of study: full-time

Murmansk




Compiled by - Elena Viktorovna Makarevich, cand. biologist. Sci., Professor, Department of Microbiology and Biochemistry, Murmansk State technical university

Guidelines for the performance of tests were reviewed and approved at a meeting of the department-developer "____" _________________ 2013, protocol No. _____.

Reviewer – Olga Yurievna Bogdanova, Ph.D. biol. Sci., Professor, Department of Microbiology and Biochemistry, Murmansk State Technical University




1. GENERAL.. 4

2. Guidelines for the implementation of control work 5

3. Questions for self-examination ... 6

6. TABLE OF OPTIONS OF CONTROL WORKS…………………….………..22

1. GENERAL PROVISIONS

The control work is one of the forms of current control of students' knowledge, and its implementation is mandatory for all students. If the work is not credited, the student should be able to correct it by re-performing the test. Students who fail the test are not allowed to take the exam.

The purpose of the test is to deepen and consolidate the knowledge of students obtained in the theoretical study of the discipline and allows the student to demonstrate the knowledge and skills acquired during the training in theory, as well as the possibility of their application in practice.

Students complete the test within the time limits set by the schedule. The performance of the control work is the final stage in the study of individual topics of the discipline "Introduction to Biotechnology".




Guidelines for the performance of control work

on the topic « Application of genetic engineering methods in biotechnology” .

To perform the control work, you must:

1. Study the theoretical data on the course;

3. Answer questions on this topic;

4. Solve provided in methodological guide test tasks;

Genetic bases for the improvement of biological objects.

Ways and methods used in obtaining more productive biological objects and biological objects with other qualities that increase the possibility of their use in industrial production (resistance to infections, growth on less deficient media, greater compliance with industrial hygiene requirements, etc.).

traditional selection methods. Variation rows. selection of spontaneous mutations. Mutagenesis and selection. Physical and chemical mutagens and their mechanism of action. Mutation classification. Problems of genetic stability of mutants based on the formation of a target biotechnological product.

Cell engineering and the use of its methods in the creation of microorganisms and plant cells - new producers of biologically active (medicinal) substances. Protoplastization and fusion (fusion) of protoplasts of microorganisms. Possibility of interspecific and intergeneric fusion. Hybrids obtained after protoplast fusion and cell regeneration. Fusion of protoplasts and production of new hybrid molecules as target products. Protoplasty and activation of "silent" genes. Possibilities of obtaining new biologically active substances due to the activation of "silent genes". Methods of cell engineering as applied to animal cells. Hybridomas. Importance of hybridomas for the production of modern diagnostic preparations.

Genetic engineering and the creation of producers of new medicinal substances using its methods. Basic principles of recombinant DNA technology. Extrachromosomal genetic elements - plasmids and their functions in microorganisms used in biotechnological processes. Basic physical and chemical characteristics of plasmids. Interaction of plasmids with the host genome. The role of plasmid and phage DNA in the genetic design of producers of biologically active substances. Transposons and their use in the construction of producers. Directed mutagenesis (in vitro) and its importance in the design of producers.

The concept of a vector in genetic engineering. Vector molecules based on plasmid and phage DNA. Chemical synthesis of DNA fragments. Sequencing methods (determining the nucleotide sequence). Chemical synthesis of a gene.

Enzymes used in genetic engineering. Restrictases. Classification and specificity. Formation of "sticky ends". Restriction enzyme E. coli R1 and the sequence of nucleotides it recognizes. Ligases and their mechanism of action.

The sequence of operations for the inclusion of a foreign gene in a vector molecule. Transfer of a vector with a foreign gene into a microbial cell.

genetic markers. Methods for identification and isolation of clones with recombinant DNA.

Problems of expression of foreign genes in microorganisms. Animal cell genes; exons, nitrons. Ensuring the expression of mammalian genes in a microbial cell. Reverse transcriptase.

Ways to overcome barriers to the expression of foreign genes. Stabilization of foreign proteins (target products) in the cell. Genetic methods providing isolation of foreign proteins into the environment.

Microorganisms of various systematic groups: yeast, eubacteria, actinomycetes, etc. as hosts in the expression of foreign genes. Specific problems of genetic engineering in the creation of new producers of protein substances, primary metabolites as target biotechnological products.

SELF-CHECK QUESTIONS

1. List the traditional breeding methods.
2. Tell us about the use of cell engineering methods in the creation of microorganisms and plant cells.
3. What are the mechanisms of action of physical and chemical mutagens?
4. Name the possibilities for obtaining new biologically active substances by activating "silent genes".
5. Expand the concept of hybrids.
6. List methods for identification and isolation of clones with recombinant DNA.
7. What is the difference between exons and introns?
8. What is foreign gene expression?

TESTS

Write the correct answer:

1. The term "reverse genetics" means the following manipulations
1. DNA - RNA - protein - protein modification - cell
2. protein - RNA - DNA - DNA modification - cell
3. RNA - RNA modification - DNA - protein
4. cell - DNA - RNA - protein - protein modification

2. Transgenic organisms are obtained by introducing a foreign gene into
1. somatic cell
2. egg
3. sperm
4. mitochondria

3. Acromegaly is characteristic of animals containing a foreign gene
1. insulin
2. interferon
3. somatostatin
4. growth hormone

4. The year when the role of nucleic acids in the transmission of hereditary information
1. 1940
2. 1944
3. 1953
4. 1957

5. The year the DNA double helix model was created
1. 1940
2. 1944
3. 1953
4. 1957

6. The first object of genetic engineering was
1 E.coli
2. S. cerevisae
3. B. subtilis

7. The first objects of genetic engineering were viruses and plasmids
1. S. cerevisae
2. B. subtilis
3 E.coli

8. As a vector for introducing a foreign gene into an animal cell, use
1. Agrobacterium plasmids
2. bacteria plasmids
4. viroids
5. SV-40 virus

9. As a vector for introducing a foreign gene into an animal cell, use
1. retroviruses
2. bacteria plasmids
3. DNA of chloroplasts and mitochondria
4. viroids

10. Do not use as a vector for introducing a foreign gene into an animal cell
1. SV-40 virus
2. retroviruses
3. Mitochondrial DNA
4. transposons
5. viroids

11. As a vector for introducing a gene into a plant cell, use
1. SV-40 virus
2. Rous sarcoma virus
3. plasmids
4. viroids

12. As a vector for introducing a gene into a plant cell, use
1. SV-40 virus
2. Rous sarcoma virus
3. Agrobacterium plasmids

13. Do not use as a vector for introducing a gene into a plant cell
1. transposons
2. DNA of chloroplasts
3. bacterial plasmids
4. viroids

14. The virus-based vector does not contain sequences responsible for
1. virulence
2. ability to replicate
3. marker sign
4. pathogenicity

15. The virus-based vector contains sequences responsible for
1. ability to transfer into the host cell
2. amplification ability
3. marker sign
4. all listed sequences

16. The vector must be
1. big
2. small
3. both statements are true

17. The use of mitochondrial and chloroplast DNA as a vector is based on
1. ring shape
2. volume
3. the presence of homologous regions with the nuclear genome
4. all statements are true

18. The number of nucleotides that make up viroids
1. 200 - 250
2. 270 - 300
3. 320 - 370
4. about 1000

19. Viroids are shaped
1. straight
2. roundabout
3. spiral

20. Transposons have a shape
1. straight
2. roundabout

21. Transposons were first discovered in
1. 30s
2. late 40s
3. 1971

22. Transposons discovered
1. Paul Berg
2. Barbara McClintock
3. Frederick Sanger

23. Year of discovery of viroids
1. 1968
2. 1971
3. 1973
4. 1977

24. Viroids are
1. 1 strand DNA
2. 1 strand RNA
3. 2 strand DNA
4. 2 strand RNA

25. Nucleic acid of viroids with protein
1. tied
2. not connected

26. Transposons play an important role in pitchfork evolution
1. yes
2. no

30. With the restriction ligation method, DNA ends are crosslinked
1. blunt-sticky
2. sticky-sticky
3. dumb-dumb

31. With the connector method, the ends of the DNA are sewn together
1. blunt-sticky
2. sticky-sticky
3. dumb-dumb

32. The use of linkers makes sense if knocking ends are formed during the destruction of 2 types of DNA by restriction enzymes.
1. namesake sticky
2. dissimilar sticky
3. stupid

33. The use of linkers makes sense if knocking ends are formed during the destruction of 2 types of DNA by restriction enzymes.
1. namesake sticky
2. blunt and sticky
3. stupid

34. Linkers are not used if the ends are formed during the destruction of 2 types of DNA by restriction enzymes.
1. namesake sticky
2. dissimilar sticky
3. stupid
4. blunt and sticky

35. Enzyme end transferase is used when sewing ends
1. namesake sticky
2. dissimilar sticky
3. stupid
4. blunt and sticky

36. Ligase is used to cross-link blunt ends of DNA in concentrations
1. insufficient
2. standard
3. redundant

37. DNA denaturation requires
1. alkaline pH
2. acidic pH
3. acidic pH and high temperature
4. alkaline pH and high temperature

38. DNA denaturation temperature (оС)
1. 37
2. 65
3. 100

39. DNA renaturation temperature (оС)
1. 37
2. 65
3. 100

40. DNA fragments pair during hybridization
1. single stranded
2. double stranded
3. single and double stranded

41. Mating is possible during hybridization
1. DNA - DNA
2. DNA - RNA
3. RNA - RNA
4. all of the above combinations

42. Hybridization of the studied nucleic acid with a DNA probe is carried out
1. in solution
2. in gel
3. based on nitrocellulose

43. Foreign DNA that has entered cells in nature, as a rule, does not show activity, as it is destroyed by the enzyme
1. ligase
2. methylase
3. restrictase
4. transcriptase

44. Birth year of genetic engineering
1. 1971
2. 1972
3. 1973
4. 1974

45. The first hybrid DNA contained DNA fragments
1. virus and bacteria
2. 2 viruses and bacteria
3. bacteria, yeast cell and virus
4. bacteria, virus and animal cell

46. ​​The first endonuclease isolated from a bacterial cell cleaved DNA molecules
1. at the place of recognition
2. at a certain distance from the place of recognition
3. in an arbitrary place from the place of recognition

47. The first restriction enzyme that cleaved a strictly defined DNA sequence was isolated
1. Meselson and Yuan
2. Meselson and Weigl
3. Smith and Wilcox

48. Polymerase contains functional domains
1. 1
2. 2
3. 3
4. 4

49. Klenov's fragment includes
1. 5'-3' polymerase and 3'-5' exonuclease
2. 5'-3' polymerase and 3'-5' polymerase
3. 5'-3' polymerase and 5'-3' exonuclease
4. 3'-5' exonuclease and 5'-3' exonuclease

50. Removes diester bond in unpaired regions of DNA
1. 5'-3' polymerase
2. 3'-5' exonuclease
3. 5'-3' exonuclease
4. 3'-5' polymerase

51. Removes diester bond in double sections of DNA
1. 5'-3' polymerase
2. 3'-5' exonuclease
3. 5'-3' exonuclease
4. 3'-5' polymerase

52. Responsible for the removal of nucleotides attached during replication
1. 5'-3' polymerase
2. 3'-5' exonuclease
3. 5'-3' exonuclease
4. 3'-5' polymerase

53. In the processes of DNA repair, cutting out oligonucleotides for a 10 bp, participates
1. 5'-3' polymerase
2. 3'-5' exonuclease
3. 5'-3' exonuclease
4. 3'-5' polymerase

54. Terminal transferase catalyzes the addition of nucleotides to the end DNA molecules
1. 5' - OH
2. 3' - OH

55. Recognize and cleave DNA molecules at arbitrary points of the nuclease
1. 1st class
2. 2 classes
3. 3 classes
4. 1st and 3rd grade
5. 2nd and 3rd grade

56. Recognize and cleave DNA molecules at a string in the recognition site or at a fixed distance from it nuclease
1. 1st class
2. 2 classes
3. 3 classes
4. 1st and 3rd grade
5. 2nd and 3rd grade

57. 1 protein in restriction endonucleases is responsible for restrictase and methylating activity
1. 1st and 3rd grade
2. 2nd and 3rd grade
3. 1st and 2nd grade
4. 2 classes
5. 3 classes

58. Different proteins in restriction endonucleases are responsible for restriction endonuclease and methylation activity.
1. 1st and 3rd grade
2. 2nd and 3rd grade
3. 1st and 2nd grade
4. 2 classes
5. 3 classes

59. False isoschizomers are
1. Hpa I and Eco RI
2. Hind III and Eco RI
3. Hpa I and Hind III

60. When DNA is run in an agarose gel, fragments will be closer to the starting line
1. short
2. long
3. short

62. To build a restriction map, it is necessary to sequentially process DNA fragments
1. 1 restriction enzyme, then 2 restriction enzyme
2. 1 restriction enzyme and a mixture of 1 and 2 restriction enzymes
3. 1 restriction enzyme, 2 restriction enzyme and their mixture

63. The first restriction map was obtained for
1. bacteriophage
2. pBR 322 plasmids
3. Rous sarcoma virus
4. SV-40 virus

64. Restriction maps allow you to determine
1. complete nucleotide sequence
2. degree of homology of DNA sections
3. violations in the work of the gene
4. gene structure

65. DNA chemical sequencing is based on
1. synthesis of a complementary DNA region
2. destruction of 1 nucleotide
3. destruction of one of the 4 nucleotides in each reaction mixture

66. DNA chemical sequencing proposed
1. Sanger and Gilbert
2. Savage and Maxum
3. Maxam and Gilbert

67. Enzymatic DNA sequencing proposed
1. Maxam
2. Gilbert
3. Sanger
4. Savage

68. During chemical sequencing, DNA is labeled
1. from one end
2. from both ends
3. full length

69. Modification of nucleotides during enzymatic sequencing involves changing the ends
1. 3'-OH
2. 5'-OH
3. 3'-OH and 5'-OH


70. In enzymatic sequencing, modified nucleotides are added compared to normal ones in
1. excess
2. equal ratio
3. lack

71. For underrestriction endonuclease is added
1. in short supply
2. excess

72. Underrestriction is usually used when using restrictases
1. coarse-grained
2. fine-chewing
3. 1st class
4. 3 classes

73. Irreversible binding of DNA to nitrocellulose requires temperature (оС)
1. 65
2. 70
3. 80
4. 100

74. Irreversible binding of DNA to nitrocellulose requires high temperature and
1. normal pressure
2. high pressure
3. low pressure
4. vacuum

75. The transfer of DNA to a nitrocellulose filter is called
1. Northern Blotting
2. Southern blotting
3. Western blotting

76. The transfer of RNA to a nitrocellulose filter is called
1. Northern Blotting
2. Southern blotting
3. Western blotting

77. The transfer of protein to a nitrocellulose filter is called
1. Northern Blotting
2. Southern blotting
3. Western blotting

78. Filter paper during blotting provides a current of buffer solution in the direction
1. electrophoresis
2. reverse electrophoresis
3. perpendicular to electrophoresis

79. The name "shotgun method" is applied to libraries
1. genomic
2. clone DNA

80. The creation of libraries begins with the synthesis of DNA on an RNA template
1. genomic
2. clone DNA

81. When creating a genomic library, the genome is represented
1. whole
2. fragmentary

82. Creating a genomic library can be considered DNA amplification
1. in vitro
2. in vivo

83. The creation of a clone library can be considered DNA amplification
1. in vitro
2. in vivo

84. Polymerase chain reaction can be considered DNA amplification
1. in vitro
2. in vivo

85. When obtaining animal proteins using a bacterial cell, it is better to use a DNA library
1. clone
2. genomic

86. The method of cell-free molecular cloning was developed in
1. 1973
2. 1976
3. 1977
4. 1985

87. Polymerase chain reaction developed
1. Berg
2. Gilbert
3. Southern
4. Mullis

88. A technique for transferring DNA to a nitrocellulose filter was developed by
1. Berg
2. Gilbert
3. Southern
4. Mullis

89. During the polymerase chain reaction, the amount of DNA increases from cycle to cycle
1. into several fragments
2. in arithmetic progression
3. exponentially

90. DNA amplification cycle in vitro takes (in minutes)
1. 5
2. 10
3. 15
4. 20

91. For the purposes of medical diagnostics, DNA amplification by means of cloning is most often used.
1. in a virus
2. in a plasmid
3. cell-free molecular

92. b-lactamase promoter
1. strong adjustable
2. weak unregulated
3.weak adjustable
4. strong unregulated

93. Promoter derived from bacteriophage l
1. strong adjustable
2. weak unregulated
3.weak adjustable
4. strong unregulated

94. Promoter derived from bacteriophage l is regulated
1. tryptophan starvation
2. lactose
3. temperature

95. The presence of introns and exons is not characteristic of DNA
1. yeast
2. plants
3. animals
4. bacteria

96. Only a eukaryotic cell is characterized by the presence
1.attenuator
2. Shine-Dalnarno sequences
3. modulator

97. Only a eukaryotic cell is characterized by the presence
1.attenuator
2. promoter
3. Amplifier

98. During transfection, ligation of the marker trait with the introduced gene
1. required
2. optional

99. The efficiency of DNA entry into cells
1. high
2. low

100. Frequency of cell DNA transformation during transfection
1. high
2. low

101. Stable transformation undergoes upon transfection 1 of
1. 10 cells
2. 100 cells
3. 1000 cells

102. The microinjection method was developed
1. Maxam and Gilbert
2. Meselson and Yuan
3. Andersen and Diakoumakos

103. Stable transformation of cells is higher with
1. transfection
2. microinjections
3. high enough in both cases

104. Microinjections transform cells (%)
1. 1
2. 10
3. 30
4. 50
5. 100

105. Promoter replicates ribosomal genes
1. Paul I
2. Pol II
3.Pol III

106. Promoter replicates structural genes of proteins
1. Paul I
2. Pol II
3.Pol III

107. Replicates genes encoding small RNA promoters
1. Paul I
2. Pol II
3.Pol III

108. For the expression of eukaryotic genes in a prokaryotic cell, it is necessary to put them under the control of regulatory elements
1. eukaryote
2. prokaryote
3. prokaryotes and eukaryotes

109. Attenuators are located between
1. 1 and 2 structural genome
2. at the end of the structural gene
3. between the promoter and the 1st structural gene
4. between the promoter and the 2nd structural gene

110. An enzyme gene is used as a marker for bacterial cells
1. thymidine kinase
2. lactose
3. Antibiotic

111. A gene is used as a marker for an animal cell
1. thymidine kinase
2. lactose
3. Antibiotic

112. With the connector method using terminal transferase, meaningless sequences are formed
1. can
2. cannot

113. The method most often used in the construction of hybrid DNA
1. restriction ligation
2. connector
3. with the use of linkers

114. In the restriction ligation method, meaningless sequences are formed
1. can
2. cannot

115. The nomenclature of restrictases was proposed
1. Smith and Nathans
2. Meselson and Yuan
3. Smith and Wilcox

116. Restriction sites with respect to 180°C rotation
1. symmetrical
2. not symmetrical

Finish sentence:

117. The _____________ method is based on the change in the permeability of the membrane when passing high-voltage pulses.
118. By sonicating aqueous emulsions of phospholipids, _______ is obtained.
119. On the formation of pores in cytoplasmic membrane based method _________
120. __________ is used to protect exogenous genetic material when it is introduced into a cell.
121. Salmon sperm DNA added to a specific gene - _________
122. Introduction of DNA using calcium precipitate - ______________
123. Regulatory sequence capable of lowering the level of transcription even in the presence of a strong ___________ promoter.
124. A double-stranded DNA fragment, necessary for polymerase to start working, is called ___________
125. Vector capable of replication in both bacterial and animal cells - _______
126. The sequence of 6-8 nucleotides responsible for the binding of RNA to the ribosome - __________ _____________.
127. The regulated promoter is called ____________.
128. The DNA sequence from which the reading of information begins - ________
129. Restriction isolated from Streptomyces albus is called _____
130. A restriction enzyme isolated from Escherichia coli is called _____
131. Restriction enzyme isolated from Streptcoccus aureus is called _____
132. Methylase isolated from Streptomyces albus is called _____
133. Methylase isolated from Escherichia coli is called _____
134. Methylase isolated from Streptcoccus aureus is called _____
135. A restriction enzyme isolated from Haemophilus parahaemolyticus is called _____
136. The enzymatic method involves the use of __________ ________
137. An enzyme responsible for the migration of certain DNA segments within a chromosome - _______________.
138. A DNA-containing virus ____________ is used as a vector for introducing genes into an animal cell.
139. Genetic elements of a cell capable of migration within a chromosome are called _____________.
140. RNA-containing viruses capable of changing the cell genome - __________.
141. In vitro construction of functionally active genetic structures is called ________________ ____________.
142. Making recombinant DNA in a test tube is called _______ _______.
143. Artificial genetic structures are called ______________.
144. Multiple doubling of a plasmid or DNA fragment - ___________.
145. The duplication of a gene in a cell or test tube is called ____________.
146. Enzyme responsible for specific labeling of DNA in a cell - _______.
147. The enzyme responsible for the restoration of the phosphodiester bond in the DNA molecule - _____________.
148. The enzyme responsible for the synthesis of the complementary DNA chain - ____________.
149. An enzyme that modifies the "blunt" ends of DNA - ______________.
150. An enzyme that introduces breaks in the double strand of DNA - ____________.
151. The enzyme __________ ____________ is responsible for the synthesis of DNA on an RNA template.
152. A restriction enzyme isolated from Bacillus subtilis is called _____
153. Promoter initiating transcription rarely - ____________
154. Promoter that often initiates transcription - _______________.
155. A small oligonucleotide containing opposite sticky ends is called ___________.
156. Protein that prevents the binding of polymerase to DNA - ____________
157. The stage of the polymerase chain reaction, when a single-stranded fragment associated with the primer is formed - _____________.
158. Introduction of DNA into cells with help. DEAE-dextran - _______________.
159. A method of introducing DNA based on a change in the permeability of the CPM by processing with electrical impulses is called




n\n	Name of textbooks, manuals and other sources	Authors (ed.)	publishing house	The year of publishing
Main:
1.	Fundamentals of biotechnology: tutorial	T. A. Egorova, S. M. Klunova, E. A. Zhivukhina	Moscow: Academy
2.	Modern biotechnology	Yeldyshev Yu.N.	M:Tydex Co.
3.	Hydromicrobiological analysis of wastewater. Methodical instructions.	Makarevich E.V. Litvinova M.Yu.	Murmansk MSTU
4.	Ecology of microorganisms	Netrusov A.I.	Moscow: Academy
	Industrial microbiology and biotechnology	Makarevich E.V.	MSTU
	Fundamentals of industrial biotechnology: textbook. allowance for universities	Biryukov, V. V.	M. : Kolos
	Molecular Biotechnology: Principles and Applications / Per. from English. N. V. Baskakova	Glick, B.	M. : Mir
	Theoretical basis biotechnologies and practical aspects of their use in the production of a number of biologically active substances from raw materials of animal, aquatic and vegetable origin in national economy Russia and medicine. Ch.1,2	Semenov, B. N.	KSTU. - Kaliningrad
	Microorganisms of fermented milk products. Methodical instructions.	Makarevich E.V., Litvinova M.Yu.	Murmansk MSTU	2009.
	Guidelines for laboratory work in the discipline "Introduction to Biotechnology"	Litvinova M.Yu.	Murmansk MSTU
Additional:
	Introduction to Biotechnology	Becker M.E.	Moscow: Food prom
	Burgey's Bacteria Key. In 2 t.	Ed. J. Holt, N. Krieg, P. Smith and others.	M.: Mir
	Industrial microbiology.	Ed. Egorova N.S.	M.: graduate School
	Technical microbiology of fish products.	Dutova E.N. and etc. .	Moscow: Food prom
	Microbiology of food production.	Verbina N.M., Kapterova Yu.V.	Moscow: Agropromizdat
	The main nutrient media for the cultivation of microorganisms. Preparation of nutrient media. Guidelines for performing laboratory work.	Bogdanova O.Yu., Makarevich E.V.	Murmansk MSTU
	Microbiology of animal products	Münch G.D., Zaupe H., Schreiter M. et al.	Moscow: Agropromizdat
	Microbiology in the food industry.	Zhvirblyanskaya A.Yu., Bakushinskaya O.A.	Moscow: Food industry




TABLE OF TASK OPTIONS

Presumed cipher digit	Last cipher digit
	1, 17, 29, 48, 134,80,127	2, 18, 31,49, 133,81,128	3, 19, 32,50, 132,82,129	4, 20, 33,51, 131,83,130	5, 21, 34,52, 130,84,131	6, 22, 35,53, 129,85,132	7, 23, 36,54, 128,86,133	8, 24, 37,55, 127,88,134	9, 25, 38,56, 126,89,135	10, 26, 30,57, 125,87,136
	11, 27, 40,52, 124,90 ,137	12, 28, 29,46, 123,91,138	13, 17, 31,45, 122,92,139	14, 18, 33,44, 121,93,140	15, 19, 40,43, 120,94,141	16, 20, 34,42, 119,95,142	1, 21, 39,41, 118,96,143	2, 22, 40,60, 117,97,144	3, 23, 30,59, 116,98,145	4, 24, 40,58, 115,99,146
	5, 25, 36,45, 114,70,147	6, 19, 26, 32, 113,71,148	7, 27, 29,51, 112,72,149	8, 28, 31,44, 111,73,150	9, 17, 37,58, 110,74,151	10, 18, 39,60, 109,75,152	11, 19, 34, 42, 108,76,153	12, 20, 30,50, 107,77,154	13, 21, 37,56, 106,78,155	14, 22, 38, 47, 105,79,156
	15, 23, 32,59, 135,43,157	16, 24, 38,49, 103,85,158	1, 25, 38,51, 102,72,159	2, 26, 58, 39, 101, 126,85	3, 27, 31,52, 100,94,127	4, 28, 37,44, 99,112,128	5, 17, 30, 56, 98,130,129	6, 18, 34, 55, 97,69,130	7, 19, 39,60, 96,123,131	8, 20, 33, 44, 95,110,132
	9, 21, 39,47, 94,127,133	10, 22, 40,51, 93,128,134	11, 23, 39, 47, 92,129,135	12, 25, 33, 44, 91,130,136	13, 26, 31,59, 90,131,137	14, 28, 30,58, 89,132,138	15, 27, 31,42, 88,133,139	16, 28, 36,45, 87,134,140	1, 17, 34, 56, 86,102,141	2, 18, 39,57, 85,103,142
	3, 19, 38,51, 84,104,143	4, 21, 38,53, 83,105,144	5, 20, 33,50, 82,106,145	6, 23, 29,57, 81,107,146	7, 22, 30,59, 80,108,147	8, 25, 29,60, 79,109,148	9, 24, 38,49, 78,110,149	10, 27, 31,54, 77,111,150	11, 25, 39,42, 76,112,151	12, 24, 34,52, 75,113,152
	13, 17, 32,41, 74,114,153	14, 18, 33,42, 73,115,154	15, 19, 40,59, 72,116,155	16, 20, 30,43, 71,117,156	1, 21, 30,44, 70,118,157	2, 22, 31,45, 69,119,158	3, 23, 29,46, 68,120,159	4, 24, 39,47, 67,121,127	5, 26, 31,60, 113,95,128	6, 27, 40,55, 66,122,129
	7, 28, 33,48, 65,123,130	8, 28, 32,49, 64,124,131	9, 17, 30,50, 73,125,132	10, 18, 38,51, 100,78,133	11, 19, 39,52, 101,77,134	12, 20, 32,52, 102,83,135	13, 21, 34,53, 103,84,136	14, 22, 29,54, 104,85,137	15, 28, 33,52, 105, 86,138	16, 23, 31,45, 106,87,139
	1, 27, 34, 55, 107,72,140	2, 26, 30,56, 108,74,141	3, 22, 40,57, 109,75,142	4, 25, 39,58, 110,76,143	5, 26, 40,59, 111,77,144	6, 17, 33,60, 112, 78,145	7, 18, 38,45, 113,79,146	8, 19, 35, 41, 114,90,147	9, 20,29,53, 115,134,	10,21,38,56,116, 133,149
	11,22,30,42,150 117,132	12, 23, 37,43, 153 118,131	13, 24, 32,44,154 119,57	14, 27, 40,68,155 120,93	15, 23, 38,66,156 121,84	16, 23, 30,51,157 122, 74	1, 22, 40,78,158 123, 59	2, 17, 39,99,159 124,55	3, 18, 33,41,160 125,96	4, 19, 29,45,143 126,87




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