Means causing inherited and fixed in the offspring directed matirovanie genome of the cells of unicellular and multicellular organisms

 

(57) Abstract:

Usage: in biotechnology, medicine, veterinary, agriculture, food industry. The inventive discovered that salts of N-substituted 3-oxypyridine can serve as a means of causing inherited and fixed in the offspring directed matirovanie genome of the cells of unicellular and multicellular organisms. 4 Il., 8 table.

The invention relates to biotechnology and may find application in medicine, veterinary medicine, agriculture and the food industry.

It is known that the beginning of 70-ies was marked by the era of genetic engineering. At the heart of modern industrial biotechnology is the possibility of a direct design of the genetic material, its introduction into living cells and realization using them contained the genetic information. The possibility of introducing any segment of DNA into cells allows you to create an industrial microorganisms, for example, to synthesize the valuable proteins. So, first described recombination DNA consisted of a DNA fragment of the virus OF and bacteriophage Pdvga1 with the galactose operon of Escherichia Coli (I. D. A. Jackson, R. H. Symons, P. Berg. Proc. Nat. Acad, Sei, USA, 1972, v.69, the ' series. From other bacteria should be noted Bacillus subtilis gram-positive, non-pathogenic, non-parasitic microorganism, as well as strains of Pseudomonas, Streptomyces, etc.

Currently, E. coli is considered melotonin producer heterolytic proteins. Moreover, E. coli cells do not have the biochemical mechanism capable of RNA splicing. Are promising filamentous fungi, yeast, insect cells and mammalian. It is possible that cells of animals in the near future can be widely used for production of recombinant proteins, monoclonal antibodies, viral vaccines, factors of immunoregulation, hormones and enzymes (A. T. Bull. Proc. 4-th Europ. Congress Biotechnol. 1987, vol 4, p. 189-202).

In genetic engineering, there are several possible approaches to the implementation of efficient expression of foreign genes in microorganisms. Ideally chemical or microbiological synthesis of eukaryotic gene or the introduction of a foreign gene inside the structural gene of bacterial or yeast cells, obtaining structural gene using reverse transcription of the corresponding mRNA that does not contain any intron from nucleotide sequence that encodes pervisory, i.e., the synthesized polypeptide is strikingly different from the normal eukarioticheskogo protein.

As for the problem of synthesis of the gene, while today it is still a formidable task, which is only slightly easier when using corresponding sets of oligonucleotides with overlapping sequences capable of self-Assembly and synthesis of dvuhtsepochnyj DNA (gene).

The second approach is a direct method of expression is based on the coupling of the structural gene in the field of translation initiation with the formation of a hybrid site recognition by the ribosome. Given that the process of removal of signal sequences in the bacterial cell is not always possible, in the design of the front of the structural gene is inserted codon of translation initiation (meinenemy codon).

According to the third scheme, the foreign gene is the first gene of the operon so that the site of initiation of translation of a foreign gene is blocked at one nucleotide site translation termination of the first gene of the operon.

Not going into detail on the intricacies of cloning of the gene and expression of the corresponding protein by methods of recombinant DNA technology, note that klonecki recipient, providing effective expression of alien/s gene/s in microorganisms refers to the second stage design. The third stage of the design it actually selection of the host cell.

The sequence of stages of work on the cloning of the gene and expression of the corresponding protein by methods of recombinant DNA technology can be represented in the following stages of design (J. Bailey. Ollis D. fundamentals of biochemical engineering. M. Mir, 1989, T. 1, S. 440-462):

obtaining fragment (alien) DNA;

the introduction of a foreign DNA fragment into the plasmid vector;

transformation;

detection of the required clone;

growing culture and isolation of plasmids;

determination of the nucleotide sequence of the cloned DNA fragment;

the design and construction of plasmids for expression;

transformation;

detection of the required clone;

growing culture and isolation of plasmids;

check nucleotide sequence of DNA;

transformation;

the cultivation of crops for the production of protein.

Thus, from the basic material we can conclude that modern genetic engineering and biotechnol is slavnosti and medicine tends to curb the development of biotechnology in General and genetic engineering in particular (Biofutur, 1989, New Sci. 1990, p. 40). In some countries (Germany, Switzerland and some others) banned the use of products of biotechnology (e.g., D. Mackenzie, New Sci. 1990, N 1707, p. 28). In genetically engineered E. coli bacteria are not only malodoznye producers heterolytic proteins, but also pose a lethal danger to humans, because they contain elements of unexpected neoplastic cell transformation.

Methods of recombinant DNA technology allows to obtain microorganisms with outstanding features, but they are malodora or unacceptable for multicellular organisms (Bayev, A. A. Genetic engineering. Journal of the all-Union chemical society them. D. I. Mendeleev, 1984, I. XXIX, No. 2, S. 4-6).

It is important to emphasize that the outstanding attributes of new strains of microorganisms are not fixed in subsequent generations. Partly this is explained, first, by the loss of the plasmid by microorganisms and, secondly, due to structural rearrangements of the genetic material of recombinant plasmids.

To obtain genetic material with useful properties are known, the application of the factors causing matirovanie genome of the cell.

Thus, the known physical factors (for example, all types of ionizing radiated).

However, the effect of different ionizing radiation is fundamentally the same: the formation of ions in the irradiated cell population is the beginning of a complex chain of physical and chemical processes leading to gross violation of biochemical reactions and, ultimately, to the emergence of gene mutations and structural rearrangements of chromosomes. Orientation processes caused by different types of ionizing radiation, unpredictable and uncontrollable, and the resulting mutations are not fixed in subsequent generations.

Known chemicals that cause mutations (ibid.).

However, the vast majority of them possess carcinogenic activity, which makes it dangerous to work with them. Moreover, mutations caused by known means, not fixed in subsequent generations.

For example, strains of E. coli have shown the possibility of directed mutation of the genome of bacteria, providing, for example, the utilization of lactose of salicin and other substrates (B. G. Hall, L. L. Parker, P. W. Betts et al. Genetics, vol. 121, No. 3, p.423; J. A. Heinemann, G. F. Sprague. -Nature, 1989, vol.340, N 6230, p.205).

But even if directed matirovanie and described for E. coli, it is not described for eukaryotes and especially for higher organisms.

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The technical result achieved when implementing the present invention, is the discovery of a group of substances with a new property, not previously described for any of the known compounds.

Such compounds according to the invention are salts of N-substituted 3-oxypyridine, which are recommended for use as a means of calling inherited and fixed in the offspring directed matirovanie genome of the cells of unicellular and multicellular organisms.

First established that there are such connections, the use of which allows you to call aimed useful mutations that are inherited and fixed in the offspring.

In Fig.1 graphically presents the dependence of the inclusion of [3H]-thymidine and [3H]-uridine (b) in the cell line carcinoma human ovarian concentration of perchlorate N-phenyl-3-oxypyridine (1), N oksifenil-3-oxypyridine (2) and N-tolyl-3-oxypyridine (3) of Fig.2,a diagram of the effect of perchlorate N-phenyl-3-oxypyridine on the growth of yeast cells of C. tropicalis strain SK-4; Fig. 2 b graphically the results of a study of the growth of the colony commercial yeast cells and populations of C. tropicalis strain CR-4 in a tough village is 2) and N-tolyl-3-oxypyridine (4); in Fig.3 diagram of the formation of the sex of the fetus when copulation reproduction of Fig. 4 schematic diagram of the effect of perchlorate N-phenyl-3-oxypyridine and N oksifenil-3-oxypyridine to switch the gene/s floor in the locus rd H or Y-chromosome evolutionary stable segments 77, 80, 97 and 124 Kb in the range of CYP1A2, is approximately 140 Kb or 0.2% of the Y-chromosome.

Example 1. Studied the effect of perchlorate N-phenyl-, N oksifenil - and N-tolyl-3-oxypyridine on tumor cell lines carcinoma human ovarian CaOThe rate of synthesis of DNA and RNA was evaluated by incorporation of [3H]-thymidine and [3H]-uridine in these cells. Evaluation of the cytotoxic action of the preparation was carried out radiometric method (Dobrynin Ya Century who issued So I. Kondratiev A. N. Problems of chemotherapy of malignant tumors. M. 1974, S. 175).

The results are shown in Fig.1 and show the dependence of the synthesis of DNA and RNA (the inclusion of [3H]-thymidine and [3H]-uridine) from the claimed compounds and their structure.

For example, under the influence of N-tolyl-3-oxypyridine twice increases DNA synthesis at the dose of 100 µg/ml (Fig.1). This effect of the drug on DNA synthesis is more niskich speed enable [3H]-thymidine. DNA synthesis increases at doses of 10.0 and 1.0 μg/ml of the indicated drug. The connection has a high cytotoxic activity in the studied doses.

The results based on the inclusion of [3H]-uridine in the cell line carcinoma of the ovary person on the concentration and structure of claimed investigational compounds allows you to select data N oksifenil-3-oxypyridine. This drug is almost completely inhibits DNA synthesis and at the same time significantly stimulates the synthesis of RNA: enable [3H]-uridine increases when a dose of 10 μg/ml and 214% relative to the benchmark. As the edge criterion activity take 50% inhibition of incorporation of [3H]-thymidine and [3H]-uridine at test concentrations of thymidine and uridine 0,510-3M

Example 2. Preparations of polytene chromosomes (HRP) from the salivary glands of Drosophila melanogaster produced by well-known methods (Kiknadze, I. I. Functional organization of chromosomes. L. Nauka, 1972, S. 212). Analysis of the functional activity of polytene chromosomes was performed according to the physiological map.

In highlighted in the last third larval age experienced drosophil F1and F5-generations nezamutnennymi plots in the third area of the rings, Balbiani (CB1) and to a lesser extent in the fourth region CB2within disk 4-3A1-6and 4-4A4-6. The other three long chromosomes contain several giant puffs with operation up to 35 drives and more, actually passing in the nucleoli. About the severity of the symptoms can be judged on the following parameters: the average diameter of polytene chromosomes experienced drosophil 2 times more than the control. The ratio of the diameter of the PUFA to the diameter of the chromosome and the length of PUFA in the experimental animals, respectively, is 1.9-2.0 and 0.5-0.8 cm as against 3.5-1,4 and 2,3-4,6 cm in the control.

PH control animals though, and contains a large number of transverse disks and areas resembling puffs, genetically few active due to the relatively large heterochromatin sites.

It is known (Kiknadze, I. I. Functional organization of chromosomes. L. Nauka, 1972, S. 212) that intensively functioning plots in the third and fourth regions CB1and CB2IV chromosomes indicate activity education ILV. In turn, the research results of the other data I-P chromosomes containing complex puffs with the functioning of 20-35 drives, essentially nucleoli, testify to intensive si).

Education excess pool rRNA (matrix, ribosomal, information and so on) in the cells detected directional matirovanie and sustainable consolidation of the new beneficial mutations in the generation confirm the possibility of occurrence in the genome more semantic information in the rRNA molecules than is encoded in the source coding DNA chain on the matrix rRNA on the mechanism of reverse transcription. Mean activation under the influence of claimed investigational compounds revertase and RNA dependent DNA polymerase, providing the reverse order of the transmission of genetic information, the synthesis of the DNA strand, complementary to the matrix and RNA.

Example 3. The study of the influence of N-phenyl-3-oxypyridine on the growth of Candida tropicalis (16 populations) was carried out according to the following procedure: in a nutrient medium (malt wort or environment Saburo) on average in 100 repeated experiments were made with varying doses of the drug (0,13333; 0,10; 0,0666; 0,03333 and 0,01333 mg/l). After 15 h of incubation at 35oC in aerobic conditions in the control and experimental groups was calculated in the camera Goryaeva the number of yeast cells (F1generation). In order to identify mutations and possible consolidation in the generation of the sign in new culture medium in the same sequence the cells/ml. After incubation, the cells are again transferred to new medium and so on, repeating this procedure with the number of repeated experiments up to 10 generations (F10).

In another experience to 0.5% increase starch solution and 2 ml of 1% aqueous solution of Baker's yeast was made 10-8mol preparation of N-phenyl - or N oksifenil-3-oxypyridine. The control variant differed from the experienced lack of study drug. After 24 h incubation at 20-25oC in aerobic conditions were counting the number of yeast cells. To identify the possible fixing mutation in the generation of the experimental and control options transferred to new starch solutions is the number of cells to match the experience 53/ml and in the control 67/ml.

The counting of yeast cells was performed in the camera Goryaeva daily. The morphology of the cells was investigated under normal biological microscope.

In Fig. 2, and presents the results of studying the influence of perchlorate N-phenyl-3-oxypyridine on the growth of yeast cells of C. tropicalis strain SK-4. The action of the claimed compounds were tested in 16 populations of cells, it was found that, on average, 100 studies of the introduction of N-phenyl-3-oxypyridine in a nutrient medium when the op is superior to the control population, incubated in wort without the addition of the drug, 2,49 times (p < 0,001). In the re-growth of new populations of experienced yeast cultures for 10 generations (time of observation) in the above culture medium without making the drug proliferation of yeast remains in average up to 1.5 times the period of F1F10generations.

Any changes in the morphology of the cells was not detected. Protein synthesis for the period F1F10generations corresponds to the speed of cell proliferation.

The effects of salts of N-phenyl - and N oksifenil-3-oxypyridine on the growth of colonies of commercial Baker's yeast on starch medium shown in Fig.2,b. A small number of Baker's yeast was inoculable in certain objects starch limiting environment. On the chart, find the exponential phase of the logarithm of the growth, cell division with constant speed. Moreover, the exponential growth phase of the Baker's yeast under the influence of N oksifenil-3-oxypyridine within 30 days is significantly higher than for control groups of phases of growth.

Insulinopenia a small number of experimental and control eukaryotes new starch environment showed a mean-oxypyridine on Wednesday, containing Baker's yeast on starch diet is found a certain period of lag phase (time of onset of cell division) with the subsequent explosive growth of colonies of cells (Fig.2,b: 3 control; 4 experience). It is important to note that under the influence of claimed investigational compounds the rate of cell division experienced eukaryotes after the lag-phase vzryvoobrazno increases, exceeding the benchmark in tens, hundreds and thousands of times, and this high rate of proliferation was fixed for a long period of time (60 days observation period).

Under normal conditions, the yeast cells are not disposed of starch (Industrial Microbiology. /Ed. by N. With.Egorova. M High school, 1989, C. 416). Significant explosion colony Baker's yeast only on starch medium indicates the directed mutation of the genome of cells in the direction of the metabolism of starch. The possibility of the synthesis of the gene (s) and fragment (s) under chemical influence on the cells creates an extremely promising field of biochemical engineering, allowing a greater degree than the method of modern genetic engineering and biotechnology, to cause the cell to synthesize new amino acid sequences, CR is of evich products.

It is known that the Cdc2 gene of yeast cells and human cells are identical and provides the normal course of events of the cell cycle. The activity of protein kinase C, encoded by the gene Cdc2, in the studied conditions, the metabolism of microorganisms, obviously, is not disturbed, as evidenced by the fact that the morphology of the yeast cells is not changed and the synthesis of proteins for the period F1F10generations corresponds to the speed of cell proliferation.

Example 3. The studies used non-pathogenic culture of E. coli and three pathogenic his serotype 026; 0119 and 0144, which were isolated from clinical material of the North-Ossetian Republican children's hospital. All the bacteria were highly resistant to the recombinants R.

The influence of claimed investigational compounds to overcome resistance highly resistant infectious recombinant microorganism, E. coli was determined in relation to the following antibiotics neomycin, monomitsina, penicillin, chlortetracycline, erythromycin, oxytetracycline, streptomycin, tetracycline, chloramphenicol.

Definition of sensitivity to antibiotics was performed according to standard techniques: diffusion in agar serial dilution in bule, then, on the surface of the seeded medium was superimposed disks at equal distance from each other and at a distance of 2 cm from the edge of the cups. Cups with disks kept 30-40 min at room temperature and further 16-18 h at 36-38oC.

Assessing the impact of perchlorate N-phenyl-, N-tolyl and N oksifenil-3-oxypyridine was carried out according to the diameter of the growth inhibition of colonies around the disk, including the diameter of the disk. The larger the zone of growth inhibition of the test culture, the higher its sensitivity to this antibiotic concentrations: 10 mm sensitivity took over sensitive. When testing drugs used 24-hour broth culture for the cultivation of which in the nutrient medium has made the following doses of antibiotics, mg/ml: 200,0; 20,0; 2,0; 0,2; 0,02; 0,002; 0,0002; 0,00002; 0,000002; 0,0000002.

In table.1 shows the results to determine the sensitivity of non-pathogenic (I) and pathogenic serotypes of E. coli 0 119977 (P), 0144886 (III) and 026028 (IV) antibiotics. From the above results show that E. coli resistant to the effects of all nine of the studied antibiotics. When introduced into the nutrient medium N-phenyl-3-oxypyridine overcome the resistance of pathogenic bacteria to neomycin, monomitsina, tetracycline, streptomycin, at the hands of Eritrean is l-3-oxypyridine serotype (II) shows high sensitivity to neomycin, the monomitsina and tetracycline, and serotype (III) to neomycin, monomitsina and penicillin. As for serotype (IV), it was not sensitive to all antibiotics.

Preparation of N-tolyl-3-oxypyridine ineffective to overcome the non-pathogenic cultures of E. coli to the effects of penicillin, chlortetracycline and oxytetracycline, but makes it a sensitive and highly sensitive to other antibiotics. Serotypes (II, IV) show different sensitivity to antibiotics under the influence of the studied drugs.

When introduced into the nutrient medium N oksifenil-3-oxypyridine serotype (IV) loses resistance to chlortetracycline, oxytetracycline, and serotype (II) to penicillin, chlortetracycline and erythromycin.

If we consider the diameter of the zones of growth inhibition of culture, under the influence of claimed investigational compounds not only overcome the resistance, but also increases the sensitivity of pathogenic and non-pathogenic cultures to antibiotics in 20-40 times that associated with blocking activity amplificating gene and, therefore, encoding glycoprotein R.

The practical significance of the obtained data was confirmed in the experiment on monkeys monkeys-rez is the playing technique of the N-phenyl-3-oxypyridine (Report of the Institute of experimental pathology and therapy of AMS USSR, 1981).

Example 4. In this work, we used more than 10,000 individuals nizamettin lines of D. melanogaster D-32 and fruit flies wild line Berlin wild. Virginia flies D. melanogaster line D-32 for mating pairs 3q 3owere placed in separate test tubes with a diameter of 18 mm at 48 hours Parent pair (PP) was removed, and their offspring were its development from egg to adult at the stern of the agar, raisins and semolina with different doses of perchlorate N-phenyl-3-oxypyridine. The control group was nurtured at the stern without the drug. From even-aged Virginia flies F1were the new family. One part PP1 breastfed at the stern with different doses of perchlorate N-phenyl-3-oxypyridine (group I - 0.05 g; group II 0.10 g; group III 0.15 g and group IV at 0.20 g per 100 g feed). Another part of PP1 was transferred to the feed without drug N. Flies derived from the last PP1, denoted as F2. A similar pattern was obtained F3and F4generations. Flies were kept in a thermostat at 25oC and every 10 days were transferred to fresh food. Every day spent fixing the duration of the development stages of time larvae (L), pupae (J), adult (s) And the number of hatched flies. The results are shown in table.2.

Study the picture of putinha spend noushich. The influence of environmental conditions, the age factor and other parameters were kept, if possible, to a minimum.

In another experience, randomly selected Virginia flies on the third day of their imaginal life was placed in pairs 3q 3oin a separate test tube for hybridization for 48 h, after which the parental couple PP was removed. A new generation of flies passed the whole cycle of development in the medium with different doses of N-phenyl-3-oxypyridine. The resulting flies were identified as F1. From virginny flies first F1generation of random selection were made new parent couples. One part PP1 transferred to normal without medication, food, and other food with the same doses as above, of the drug. From F1in the same way depending on the content on normal diet (N) or at the stern with the product (A) obtained F2-generation adults, and from F2then F3F5-generations. Every 7-8 days, flies were transferred to fresh food. In all variants of the experiment was used Virginie day flies, which were kept in a thermostat at a temperature of 200,1o; 250,1oand 300,1oC. Accounting of dead flies were daily. The results are shown in table.2.

The minimum and maximum is at 20oand 30o22 1,0 1,5 and 10 days, respectively (P < 0,001). In this work, we used the results observed fundamental convergence of several repeated experiments.

In table. 3 shows the results of studying the influence of perchlorate N-phenyl-3-oxypyridine for the duration of the development stages of Drosophila discomycetes line D-32. It may be noted that in the F3and F4generations is almost a complete synchronization of the average time of development of the larval stage of pupation and adult in control and experimental flies. The density of F1populations from the PP, only once fed the drug, in terms of N content (aft without drug) increases relative to the control experiment twice, and this high fertility is inherited from F1F5generations (observation period).

In table.2 shows the results of rearing pairs of D. melanogaster wild population at the stern with N-phenyl-3-oxypyridine. Presented in table. 2 data suggest that under extreme conditions (20oC or 30oC), i.e. in conditions where protein synthesis in addition to heat shock protein in the organism of Drosophila almost completely stopped, there are strong F1generation p is 10% population density has increased 2-4 times; average and maximum lifespan increased by 110-160% relative to the targets fixed in the F1F5generations (time of observation). The experiment will explain from the point of view of the influence of claimed investigational compounds on the signal portions of genes ontogeny of Drosophila.

Example 5. In this work, we used nonlinear rats, rats lines Wistar and mice, as well as nizamettin Drosophila lines D-32. To study the effect of salts of N-phenyl - and N-oxoferin-oxypyridine on generative activity and the possibility of artificial regulation of the sex ratio of animals were selected on the principle of steam-analogues with regard to age and body weight. In the experience used 70 Mature rats of both sexes. All animals were divided into three groups in pairs (male female). Composed of parent pairs were otsuzhivalis in a separate cell, where he remained during the whole experiment (to get these pairs of five generations of offspring) in the same conditions of normal feeding, water regime and content in vivarium. Animals of the control group (1) in the amount of ten pairs of "families" were mating of males and females six months of age, weighing 180-220 g Animals in the second group, cocoadialog N-phenyl-3-oxypyridine by oral administration via a probe in the form of alcohol solution in doses of 10 mg daily 1 time per day for 15 days. In the third group of eight pairs of parents of similar age and weight, and that in groups I and II, male for 38-40 days before mating was nurtured study drug at the same dose. Experiments were performed for the control and experimental groups of animals in April, may, September, when the sex ratio according to literature data (p.p. Gambaryan Dulsky N. M. A rat. M. the Soviet science, 1955, S. 252) is male-female, 50/50; 42,2/57,0; 45,0/55,0 respectively.

The effects of N-phenyl-3-oxypyridine at puberty, the emergence of pripadov, weight of testes and epididymis of rats, and the ratio of the mass of bodies to the body weight and the mixing ratio of the sexes is shown in table.4-8. From the above table.4 results we can conclude that the rat pups born from feeding the drug of parents, as in group II and group III revealed the true acceleration of puberty in 38-26 days, the time of occurrence of the first pripadov reduced from 116-118 days in control to 89-80 days in the experimental groups, i.e., 27-26 days.

The accounting issue in "families" were within 92 days after the first litter, as this period is the average time interval between problemami the control rats (table.4). It should be noted that in the work of palsolidarity increase in the quantity peculiar to each species (Veterinary encyclopedia, so 4, S. 1045, 1973).

For a specified period of ten test "families" (group I) litter amounted to 119 of rats, whereas in ten experienced "families" (group II) 239 rat in eight "families" group III 203 rat, five "families" group IV 56 of rats. In terms of one pair of the number of pups is: group I - 12; group II, 24; group III 25; group IV 11 of rats. From the above table. 4 data suggest that population growth is not due to the increase of individuals in the offspring, and by reducing the intervals between problemami and increase the number of pripadov: group I (control) average time intervals between problemami is 92 days (range 90-96 days), group II 34 days (range 27-45 days), group III 36 days (range 28-49 days) and group IV 94 days (range 90-99 days). Data on fertility and sex composition of offspring rats consumed the drug is presented in table.5 and 6.

The increase in 2 times and more fertility nonlinear rats by reducing intervals between problemami, and the emergence of early puberty under the influence of N-phenyl-3-oxypyridine watched for five generations.

During the observation period is not marked by notable starane the CA postnatal life did not differ from control: within 24 h after birth weight of rats in the control families were 6-8 g; in group II, 6-7 g; group III 7-9 g; group IV 7-9 g

Weight gain, the opening of the eyes, the acquisition of a wool cover in the rats in the experimental families are in accordance with the physiological norms.

Analysis of metaphase plates of bone marrow in rats showed that chromosomal aberrations are absent, all acrocentric chromosomes, normal, spiralization not broken.

The experiment proved the existence of a linear relationship between the effect of N-phenyl-3-oxypyridine not activation of the testicles and their appendages, resulting in rapid and intensive growth of their mass is greater than the weight of testes and epididymis in the control rats of the same age and weight (table.7), and shift the sex ratio in rats towards males (table.6).

Histological examination of the testes of rats, stained Felgen and brush, showed a significant increase in the thickness of the walls of the convoluted tubules of the testes to 250-320 μm (control 150-200 μm), the increase of all layers of the spermatogenic epithelium, as well as the number of actively motile spermatozoa (table.8).

Leydig cells that produce male sex hormones and supports spermatogenesis, functionally active, resulting in obeidallah, detected in control rats (table.8).

Example 6. In table.9 presents results on the effect of N-oxiranyl-3-oxypyridine on ontogeny and sex ratio in D. melanogaster discomycetes line D-32.

As you know (Harmful substances in industry. L. Chemistry, I. 1, 1976), salts of metaphosphoric acid cause a disruption in metabolic processes. However, in contrast settled in physiology concepts (D. C. Page, R. Mosher, E. U. Simpson, et al, Cell, 1987, vol 51, p.1091) under the influence of N4PO3sex ratio in Drosophila F1generations is shifted in the direction of females. Although F1the generation of the ratio q/othe experience was 1.06:1,0 against 1,30:1.0 in control. Dual action metaphosphate sodium and N oksifenil-3-oxypyridine leads to a higher frequency splitting females 1,60: 1,0.

Thus, the above in examples 5 and 6 the experimental results clearly show that depending on the nature of the influence of salts of N-aryl-3-oxypyridine on the generative functions, for example rats and Drosophila is different activation of growth factors primary sexual characteristics encoded by the X - and Y-chromosomes, therefore, different and the frequency splitting of the floor towards musciano, in embryogenesis differences in the growth rate of the embryos, male and female, are determined by the Y-chromosome and, apparently, factor in the determination of the testes (Fig.4).

The analysis presented in the sample data indicates a high efficiency of the claimed compounds as substances that cause inherited and fixed in the offspring directed matirovanie genome of the cells of unicellular and multicellular organisms.

For the first time revealed the effect of the claimed compounds in the host cell genome, the author gave the name to the client morphogenesis.

Directed matirovanie and the inheritance of a new mutation in greater speed is achieved cooperationin (sexual) way, which follows from the examples of the action of salts of N-aryl-3-oxypyridine on various strains of E. coli, yeast, fruit flies and rats. The obtained experimental results confirm the assumption that investigated the claimed compounds significantly affect the processes of "editing", i.e. the appearance of greater genetic information in RNA molecules than is encoded in the original DNA. Thereby is ensured by the mechanism (reverse common biological dogma) transmission distribution of gene activity is underwater generation of cells to another during embryogenesis (Express morphological inheritance of the trait).

Express morphogenesis in contrast to modern genetic engineering and biotechnology allows for the use of the claimed compounds to modify the genome of the unicellular and multicellular organisms and, obviously, allows to realize the huge potential evolution in humans and animals. To them in the first place include the possibility of significant bias of the active phase of human activity up to 100 years and more successful fight against diseases such as AIDS, cancer, cardiology, infectious diseases associated with dysfunction of the endocrine system, creating new outstanding strains of unicellular and multicellular organisms, the fixation of beneficial mutations in the generation, etc.

Express morphogenesis free from those drawbacks, which are characteristic of modern genetic engineering and biotechnology. At the same time, in genetic engineering, we have a paradoxical situation, when medicines biotechnology for medicine, agriculture and other areas relatively ineffective in practice (monoclonal antibodies, hematopoietic growth factors, interferons, protein hormones and so on). World merchandise Reny company Business Communications, USA). It is expected that the share of the world market of medicines have 14,086 billion. According to forecasts of economists research firm SRI International, USA global sales of therapeutic drugs, diagnostics, vaccines, pesticides and medicines for agriculture in 2000 will be about 50 billion dollars. from the total product sales of the latest biotechnology volume 66 billion.

The calculations of the development of the world market of products of modern genetic engineering and biotechnology them at a very low efficacy and high toxicity confirm the high efficiency of the introduction of rapid morphogenesis in practice. Possibly Express morphogenesis will become the leader of the Foundation, which will develop the global biotechnology industry of the XXI century (the idea of competition for Japanese-American-European countries), which was proposed by the firm Cetus,USA and supported by the Japanese firm Suntory Ltd.

The use of salts of N-substituted 3-oxypyridine as a means of calling inherited and fixed in the offspring directed matirovanie genome of the cells of unicellular and multicellular organisms.

 

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(2) -NH-(CH2)SO2-NH-R5where R5hydrogen, alkyl, or CHp=1 or 2;

(3) -NX-R6where the X group is-CH - or a nitrogen atom; R6group-СОR7where R7alkyl or alkoxy, or a group-0-C0-NH-R8where R8alkyl;

(4) -NH< / BR>
(5) -NH-(CH2)3-OR10where R10alkyl;

(6) -NH-(CH2)10-NH-CO-NR11R12where R11and R12lower alkyl;

(7) -NH-CHNO

(9) -NH-(CH2)5-0-(CH2)5-0-(CH2)5-H

(10) -NH-(CH2)3-0-CO-NH-R14where R14alkyl;

(11) -NH-CH< / BR>
(12)CHwhere m=0 or from 1 to 6; R9, R15and R16the same or different and represent hydrogen or alkoxygroup, provided that when k=0, group a sure formula

FIELD: corrosion protection.

SUBSTANCE: invention, in particular, relates to protection of oil-field equipment to suppress vitality of microorganisms and to inhibit corrosion in hydrogen sulfide-containing and acidic media, in oil production, transportation, and storage systems as well as in flooded formations. Task is solved by that, in a method of preparing corrosion inhibitor-bactericide, alkyl-substituted pyridines are brought into reaction alkyl bromides at elevated temperature, said alkyl-substituted pyridines being picolines or picoline fractions at molar ratio of picolines or picoline fractions to C10-C16-alkyl bromides 1.05:1. Picolines or picoline fractions are introduced into three-step reaction each time by 0.35 mole with time interval 40-50 min, while overall reaction proceeds for 2-5 h at 120-140°C. If necessary, reaction product is mixed with solvent to form 20-70% reagent solution.

EFFECT: simplified preparation technology and imparted hydrogen sulfide bacteria growth suppression.

4 tbl, 10 ex

Ionic liquids ii // 2272043

FIELD: organic chemistry.

SUBSTANCE: invention relates to new ionic liquids designated for using in electrochemical cells and in organic synthesis. Invention describes ionic liquids of the general formula: K+A- (I) wherein K+ represents one of cations of the group consisting of the following formulae: wherein R1-R5 can be similar or different and can be bound to one another by a simple or double bond also, and each of them separately or in common can represent the following values: hydrogen atom (H), halogen atom, (C1-C8)-alkyl radical that can be partially or completely substituted with the following groups but preferably with fluorine atom (F), chlorine atom (Cl), N-[CnF(2n+1-x)Hx]2, O-[CnF(2n+1-x)Hx], SO2-[CnF(2n+1-x)Hx] or CnF(2n+1-x)Hx wherein 1 < n < 6 and 0 < x < 2n+1; A- means anion taken among the group consisting of [PFx(CyF(2y+1-z)Hz)6-x]- wherein 1 ≤ x ≤ 6, 1 ≤ y ≤ 8 and 0 ≤ z ≤ 2y+1. Invention provides the development of ionic liquids showing broad range of liquid state, high thermal resistance and low corrosive activity.

EFFECT: improved and valuable properties of ionic liquids.

3 ex

FIELD: organic synthesis.

SUBSTANCE: invention relates to technology of preparing pyridinium hexafluorophosphate, which is convenient intermediate for synthesis of lithium hexafluorophosphate used as ionic electrolyte component for lithium chemical power sources. Pyridinium hexafluorophosphate is prepared via reaction phosphorus pentachloride with fluorination agent such as ammonium hydrodifluoride followed by treatment of resulting intermediate with pyridinium salt solution.

EFFECT: avoided use of dangerous and inconvenient liquid anhydrous hydrogen fluoride and use of extreme temperature parameters.

2 ex

FIELD: organic chemistry, chemical technology.

SUBSTANCE: invention relates to a method for preparing N-phenylpicolinium salts and phenyl-substituted picolines labeled with tritium by phenyl ring of the general formula: [CH3C5H4N+C6H*5]BF-4 and CH3C6H*5C5H3N. Method involves arylation of 2-, 3- and 4-picolines with free phenyl cations obtained in beta-decay of tritium in composition of two-fold labeled benzene in the presence of potassium tetrafluoroborate in the closed system. Method provides improving method of synthesis of labeled benzene and to use ion-molecular reaction of tritium-labeled phenyl cations for simultaneous, simple and a single-step synthesis of N-phenylpicolinium salts and phenyl-substituted picolines labeled with tritium that can be used in investigations of metabolism of biologically active heterocyclic derivatives.

EFFECT: improved method of synthesis.

1 dwg, 1 ex

FIELD: production of alkoxy-(alkyl-substituted) methylpyridin chlorides of branched structure used as emulsifying agents, solubilizing agents, detergents, disinfectants, auxiliary textile products.

SUBSTANCE: proposed method includes alkylation of pyridin by alkyl-substituted chloromethyl ethers which are obtained through interaction of three starting components: higher alcohols or their fractions, carbonyl compound which is just aldehyde or ketone and chlorinating agent containing sodium chloride and sulfuric acid.

EFFECT: facilitated technology; enhanced economical efficiency and safety.

4 cl, 7 ex

FIELD: chemistry.

SUBSTANCE: present invention concerns the salts containing bis(trifluoromethyl)imide anions and saturated, partially or completely unsaturated heterocyclic cations, method of production and application thereof as ionic liquids.

EFFECT: production of new salts to be used as ionic liquids.

19 cl, 5 ex

FIELD: chemistry.

SUBSTANCE: claimed invention relates to method of obtaining organic salts, which contain anions of bis(perfluoroalkyl)phosphinate and can be applied in organic synthesis. Difference of claimed method lies in the fact that it includes carrying out reaction of tris(perfluoroalkyl)phosphinoxide with alcohol and organic base, stronger than alcohol.

EFFECT: elaboration of new method of obtaining organic salts with properties of ionic liquids.

11 cl, 14 ex

FIELD: chemistry.

SUBSTANCE: invention relates to an agent, which is in form of fluorinated 1,4-naphthoquinone derivatives of general formula (I) which have cytotoxic effect on human cancer cells in a culture. In general formula (I) 1) R1=NHC(CH3)3, R2, R3=F; 2) R1=NHCH2CH2SCH3, R2, R3=F; 3) R1=N(CH2CH2)2, R2, R3=F; 4) R1=N(CH2CH2)2, R2, R3=F; 5) R1=NHCH2CH2CH2CH3, R2, R3=F; 6) R1=NHC6H5R2, R3=F; 7) R1=H(CH3)CH2CH2OH, R2, R3=F; 8) R1, R3=NHCH2CH2CH2CH3, R2=F; 9) R1=N(CH2CH2OH)2, R2, R3=F; 10) R1=NHC6H5, R2=CH3, R3=F; 11) R1=OCH3, R2, R3=F; 12) R1=NH(CH2)2SS(CH2)2NH(2-pentafluoro-1,4-naphthoqunonyl), R2, R3=F; 13) R1=NHC2H5, R2, R3=F; 14) R1=N+C5H5, R2=O; R3=F; 15) R1=NHCH2CH2OH, R2,R3=F; 16)R1, R2=OCH3, R3=F.

EFFECT: proposed compounds can be used in medicine as a base for designing drug formulations of preparations used in malignant growth therapy.

2 dwg, 4 tbl, 3 ex

FIELD: chemistry.

SUBSTANCE: invention relates to a novel chemical compound, specifically to rac-N-[2,3-di(tetradecyloxy)prop-1-yl]pyridinium bromide, which can deliver nucleic acids into mammal cells: .

EFFECT: compound has low toxicity and in form of an alcohol solution, the compound can deliver nucleic acids into mammal cells.

1 cl, 5 ex, 4 tbl, 1 dwg

FIELD: chemistry.

SUBSTANCE: described is a method of preparing spherical particles of solid triphenyl boron-pyridine (TPBP) by separately feeding into an intensely stirred reaction zone (i) a stream of pyridine, and (ii) a stream containing a solution of a sodium hydroxide-triphenyl-boron adduct (TPBA), as a result of which total concentration of TPBA in the merged streams ranges from 1 wt % to 6 wt %, and simultaneously removing the product stream from said reaction zone and extracting TPBP particles.

EFFECT: method can be used on an industrial scale.

4 cl, 9 ex, 2 dwg

FIELD: plant growing, in particular, growing of plants through irradiation of one plants by means of other plants.

SUBSTANCE: apparatus has chamber made in the form of body having section defined by two similar ellipses. Said ellipses intersect one another so that one of focuses is single for both the ellipses. Two volumes of radiation transmitting material are provided within said chamber, said volumes being adapted for placing of seeds-emitters and seeds-receivers. One of volumes is made spherical and is located within focus single for both the ellipses and other volume is made in the form of toroid extending along line of second focuses of said ellipses. Shape and arrangement of volumes made from radiation transmitting material allow capability of irradiation of seeds to be increased. Also, seeds to be irradiated may be treated using number of kinds of seeds-emitters.

EFFECT: wider operational capabilities and increased efficiency of apparatus.

1 dwg

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