RU2492473C2 - Method of biotesting on seed germination - Google Patents

Abstract

FIELD: biotechnology.

SUBSTANCE: uniform stacking of seeds of round red radish with white tip is carried out on the filter paper in a Petri dish. 5 ml test sample of water is poured into the dish, and the liquid level in the dish must be lower than the surface of the seeds. At that the filter paper is taken of round shape with the diameter of the Petri dish with the mark on the edge, a pattern with holes for marking the places of planting seeds is placed on it. And one of the holes is oriented relative to the filter paper mark. Then the marked filter paper is placed in a Petri dish with the orientation of its mark in a northern direction on the compass, then the seeds are laid in the Petri dish on the marked places of planting on the filter paper. The water under study is poured, after germination of seeds prior to taking seedlings from the Petri dish for measuring the length of the root in each seedling, the angle of direction of its root is measured by compass as a factor of the azimuth of the root. In undergerminated seeds the azimuth of the root is taken as the absence of a quantitative value and a dash is put in the log of measurements. Subsequently, in all the seedlings in a Petri dish the length of the root is measured, and this length in the undergerminated seeds is taken as zero.

EFFECT: improvement of the method.

15 cl, 5 tbl, 1 ex

Description

The invention relates to environmental engineering and can be used in monitoring the quality of water samples of rivers and ponds by testing the growth of the roots of various types of test plants, as well as testing filtrates from various landfills and solid waste storage facilities.

A known method of testing water pollution by the time of growth of plant roots (Pat. 2402765 Russian Federation, IPC G01N 33/18. Method of testing water pollution by the time of growth of plant roots / Mazurkin P.M., Evdokimova O.Yu. (Russian Federation); applicant and patent holder of the Mari State Technical University - No. 2009133898/04; filed 09.09.2009; publ. 10.27.10), in which, before the main testing experiments, an additional experiment is carried out to determine the rational period of germination of the seeds of the test plant, at the same time, for each type of plant, their term is determined ionic germination of seeds.

The disadvantage is that the method takes into account only the influence of the time of germination of seeds in the Petri dish on the growth of the roots of the plant, when in the Petri dish the growing roots of 50 seeds begin to interfere with each other.

There is also a method of testing water pollution by the growth of plant roots in accordance with the biotesting procedure for seed germination (Appendix 10. Sanitary rules and norms SanPiN 2.1.7.573-96 "Hygienic requirements for the use of wastewater and their rainfall for irrigation and fertilizer" (approved. Decree of the State Committee for Sanitary and Epidemiological Supervision of the Russian Federation dated October 31, 1996 No. 46), including the uniform placement of 30 or 50 pieces of red round radish seeds with a white tip or white mustard (Sinapis alba) on filter paper in a 10 cm diameter Petri dish (Consolidated report e CMEA member countries on March 7, 2005. Budapest, 1975, pp. 2-4) Moreover, 5 ml of investigated and pure water are poured into each Petri dish at 4-8-fold repetition, while the liquid level in the cups should be lower than the surface of the seeds, then the cups are covered and placed in a thermostat at a temperature of 20 ° C. And in the absence of a thermostat, an experiment is possible in room conditions, but then, due to temperature fluctuations, it is difficult to compare the results carried out at different times. Before use, Petri dishes are sterilized in an autoclave at 2 atm for 10 minutes or in boiling water for 30 minutes. The experiment ends after 72 hours. The length of the roots is measured, excluding from the series of data the five smallest values, including not sprouted seeds. If, in comparison with the control, the seeds in the test water did not germinate at all, or the root length as a percentage of the control is below 70%, then the test water is unsuitable for irrigation. The threshold of 70% is justified by the fact that the soil, due to sorption ability, reduces the inhibitory effect of the studied water. With the root length in the experiment over 120% of the control, it is assumed that water has stimulating properties. Note: The test for seed germination can be carried out with the seeds of other plants and, first of all, plants that are planned to be grown under irrigation.

The disadvantage is that the method does not take into account the influence of cardinal points on fluctuations in the wave disturbance of plants along the length of the roots, when the distribution of the length of the roots is related to the azimuth of the planting of seeds, the regional features of the water body for testing the seeds of other plants are not taken into account. Moreover, 50 seedlings after 72 hours are intertwined with each other, not only interfering with the measurements, but also forming mutual strong intraspecific competition with the development and growth of roots. Pi non-sprouted seeds are not taken into account in the measurements of the prototype.

The technical result is to increase the accuracy of testing water samples by plant seeds by achieving optimal conditions for root growth of each plant, taking into account the seeds of the test plant in measurements, and also reducing the complexity of the measurements by reducing the number of seeds from 50 to 12, that is, more than four times, and optimizing the number of repeats of the Petri dish for the main experiments for each water body due to preliminary experiments.

This technical result is achieved by the fact that the method of biotesting for seed germination, including uniformly placing 30 or 50 seeds of red round radish with a white tip on filter paper in a Petri dish, into which 5 ml of the test or pure water sample is poured, while the liquid level in the cup should be below the surface of the seeds, then the cup is covered with a lid, after 72 hours of germination, the root length of each seedling is measured, and the measurements are carried out for seedlings, excluding the five lowest values from the data series, than the test for seed germination can be carried out with the seeds of other plants and, first of all, plants that are planned to be grown under irrigation, characterized in that the filter paper is adopted in a round shape in diameter of a Petri dish with a mark on the edge, a template with holes is laid on it for marking the places of planting seeds, one of the holes being oriented relative to the label of the filter paper, then the marked filter paper is placed in a Petri dish with the orientation of its label in the north direction along the compass, etc. which is similarly done with other Petri dishes in repeats of the experiment, then seeds are laid on filter paper without marked pattern on each marked Petri dish, the test water is poured, after germination of the seeds before measuring the length of the root, each seedling measures its direction angle as an indicator of the azimuth of the root moreover, the root azimuth of non-germinated seeds is taken as the absence of a quantitative value and a dash is put in the measurement log, in the future, the length of the root is measured for all seedlings, and this d the root inu is measured for all seeds, including non-germinated seeds, in which the root length is taken to be zero, after testing the test water and measuring the length and azimuth of the roots, statistical modeling reveals the general biotechnical pattern of the distribution of root lengths from the azimuth of seed planting for each Petri dish and their combination, and subsequently calculate the correlation coefficient for all samples of the studied water and for their combination, then compare them with a sample of control water, and for testers Seeds of seeds of other plants, and, first of all, plants that are planned to be grown under irrigation, first conduct regional and basic experiments for each water body.

Identification by the method of identification of biotechnological patterns of change in the root length of each plant sprout, depending on the azimuth of planting seedlings for 72 hours of seed germination, is performed according to the generalized formula:

L = L ₁ + L ₂ ,

L ₁ = L ₀ , L ₂ = Acos (πφ / p ± a ₇ ),

, $$p=a_4\pm a_5{\varphi }^{a_6}$$

, $$A=a_{\mathrm{one}}exp(-a_2{\varphi }^{a_3})$$

, $$p=a_{\mathrm{four}}\pm a_5{\varphi }^{a_6}$$

,

where L is the root length of each plant sprout in the Petri dish, mm;

L ₁ - the first component, taking into account the effect of pollution in its pure form, mm;

L ₀ - theoretical independent value of the root length from the azimuth of seed planting, shows that this value is dependent on other factors, mm;

L ₂ - the second component of the vibrational disturbance of the plant

the root length of each sprout in the Petri dish for 72 hours, characterizing the influence of the side of the world, mm;

A - half the amplitude of the vibrational disturbance of the roots of seedlings, mm;

p - half the period of oscillatory changes in azimuth of seed planting, degrees .;

φ is the azimuth of seed planting, deg .;

and ₁ ... a ₇ are the parameters of the finished statistical model, the model parameters take specific values for specific conditions of seed germination with the test water for irrigation.

The remaining patterns are still not clear enough on the meaningful interpretation.

The essence of the technical solution lies in the fact that the filter paper is made round in the diameter of the Petri dish with a mark on the edge, a template with holes for marking the seed planting places is laid on it, one of the holes being oriented relative to the mark of the filter paper, with the mark of the seed planting places applied with a pencil in the form of dots, after applying the marking, the template is removed, the marked filter paper is placed in a Petri dish with the orientation of its label in the north direction along the compass.

The essence of the technical solution lies in the fact that the template with holes for marking the places of planting seeds is made in the form of a circle along the diameter of the Petri dish, and the holes for marking the places of planting seeds are clockwise through 30 degrees, while the distance between the holes and the center of the template is approximately 30 mm

The essence of the technical solution lies in the fact that the length of the root is measured for all seedlings, and this root length is measured for all seeds, including non-germinated seeds, for which the root length is taken to be zero, and the length of the seedlings is measured sequentially, starting from the first seedling along the way clockwise from 30 to 360 degrees.

The essence of the technical solution also lies in the fact that in a preliminary experiment, when comparing with a control sample of pure water, the seeds in the studied polluted water did not germinate at all, or the root length as a percentage of the control is below 70, then the water is not suitable for irrigation, and according to the prototype the threshold of 70% is justified by the fact that the soil, due to sorption ability, reduces the inhibitory effect of the studied water, and when the root length in the experiment exceeds 120% of the control, it is assumed that the water has stimulating properties, then for substantiating the number of repeats of Petri dishes in the main experiments, perform a correlation analysis of the seedlings in each Petri dish and from their totality from the preliminary experiment, calculating the statistical correlation coefficient.

The essence of the technical solution also lies in the fact that for river water intake in the city of Yoshkar-Ola, the main experiments on germinating red round radish seeds with a white tip include uniform seed placement in a Petri dish in the amount of 12 pieces in place after 30 degrees of planting azimuth in four repetitions Petri dishes.

A positive effect is achieved by determining the conditions for the most favorable growth of the roots of the test plant in order to increase the accuracy of testing water pollution, especially natural and other contaminated water of various quality, which is planned to be used for irrigation of crops.

The novelty of the technical solution lies in the fact that the experiment on seed germination is performed by changing the root length of the test plant, depending on the azimuth of planting the seed and the azimuth of the root of each seedling.

The proposed technical solution has significant features, novelty and a significant positive effect. We have not found any materials discrediting the novelty of the technical solution.

Figure 1 shows the layout of 12 seeds of red round radish with a white tip in a Petri dish after 30 degrees (numbers indicate the places of planting seeds); figure 2 shows the angle of determination of the azimuth of the landing φ and the azimuth of the root of the seedlings φ _to ; figure 3 presents the image when determining the azimuth of the root of the seedlings φ _to ; figure 4 shows a graph of the first component of the distribution of the root length of 12 seeds of red round radish with a white tip from the bearing azimuth with growth for 72 hours in the first Petri dish; figure 5 shows a graph of the second component of the oscillatory disturbance of the plant along the length of the root of each seedling in the Petri dish depending on the azimuth of the seed planting; figure 6 shows a General graph for a two-term mathematical model in the first Petri dish; 7 - the same in Fig.6 in the second Petri dish; in Fig.8 - the same in Fig.6 in the third Petri dish; in Fig.9 - the same in Fig.6 in the fourth Petri dish; figure 10 is the same in figure 6 in the fifth Petri dish; in Fig.11 - the same in Fig.6 in the sixth Petri dish; in Fig.12 - the same in Fig.6 in the seventh Petri dish.

The method of biotesting for germination of seeds in the General case includes the following steps.

A water sample is taken, its conservation and preparation is carried out for each test.

In the laboratory, a seed germination test is performed. To do this, the filter paper is assumed to be round in shape with a diameter of 10 cm Petri dish with a mark on the edge, a template with holes for marking the seed planting places is laid on it, one of the holes being oriented relative to the filter paper mark, with the seed marking being applied with a pencil in the form points.

The template is made in the form of a circle along the diameter of the Petri dish, and the holes for marking the seed seats are placed clockwise after 30 degrees, while the distance between the holes and the center of the template is about 30 mm.

After marking, the template is removed, and the marked filter paper is placed in a Petri dish with the orientation of its label in the north direction along the compass. Do the same with other Petri dishes for all repetitions of the experiment. Moreover, before use, Petri dishes are sterilized in an autoclave at 2 atm for 10 minutes or in boiling water for 30 minutes.

Then, in each Petri dish, at the marked places of planting, seeds of the test plant are placed on filter paper. Then, 5 ml of the studied water sample is poured into each Petri dish at 4-8-fold repetition, while the liquid level in the cups should be lower than the surface of the seeds. Next, the cup is covered with a lid and placed in a thermostat at a temperature of 20 ° C, and in the absence of a thermostat, an experiment is possible in room conditions, but then, due to temperature fluctuations, it is difficult to compare the results carried out at different times.

After 72 hours of seed germination before measuring the root length, each seedling measures the angle of its direction as an indicator of the azimuth of the root, moreover, the seed azimuth is taken for non-germinated seeds for a quantitative value and a dash is put in the measurement log.

In the future, the root length is measured for all seedlings, and this root length is measured for all seeds, including non-germinated seeds, for which the root length is taken to be zero, and the length of the seedlings is measured sequentially, starting from the first seedling clockwise from 30 to 360 degrees.

After completing the test of the test water and taking measurements of the length and azimuth of the roots, the general biotechnical regularity of the distribution of the length of the roots from the azimuth of planting seeds for each Petri dish and their combination is revealed by statistical modeling.

Identification by the method of identification of biotechnological patterns of change in the root length of each plant sprout, depending on the azimuth of planting seedlings for 72 hours of seed germination, is performed according to the generalized formula:

L = L ₁ + L ₂ ,

L ₁ = L ₀ , L ₂ = A cos (πφ / p ± a ₇ ),

, $$p=a_4\pm a_5{\varphi }^{a_6}$$

, $$A=a_{\mathrm{one}}exp(-a_2{\varphi }^{a_3})$$

, $$p=a_{\mathrm{four}}\pm a_5{\varphi }^{a_6}$$

,

where L is the root length of each plant sprout in the Petri dish, mm;

L ₁ - the first component, taking into account the effect of pollution in its pure form, mm;

L ₀ - theoretical independent value of the root length from the azimuth of seed planting, shows that this value is dependent on other factors, mm;

L ₂ - the second component of the vibrational disturbance of the plant along the root length of each seedling in the Petri dish for 72 hours, characterizing the influence of the side of the world, mm;

A - half the amplitude of the vibrational disturbance of the roots of seedlings, mm;

p - half the period of oscillatory changes in azimuth of seed planting, degrees .;

φ is the azimuth of seed planting, deg .;

a ₁ ... a ₇ - parameters of the finished statistical model, the parameters of the model take specific values for specific conditions of germination of seeds with test water for irrigation.

After identifying the biotechnological pattern of changes in the length of the root of the plant, depending on the azimuth of planting, the significance of each component of the statistical model is determined.

According to the adaptability coefficient, equal to the ratio of the wave second component to the first component of the model, when this coefficient is less than unity, it is estimated that the adaptability of plants to irrigated water is doubtful, and if the unit is exceeded, it is quite reliable.

Then, the correlation coefficient is calculated for all samples of the studied water and for their combination, and compared with the control water sample, and for testing seeds of other plants and, first of all, plants that are planned to be grown under irrigation, they first conduct regional tests for each water body preliminary and main experiments.

In a preliminary experiment, when comparing with a control sample of pure water, the seeds in the polluted water under study did not germinate at all or the root length as a percentage of the control was below 70, then the water is unsuitable for irrigation, and the prototype threshold is 70% justified by the fact that the soil, thanks to sorption ability, reduces the inhibitory effect of the studied water, and with the root length in the experiment over 120% of the control it is assumed that the water has stimulating properties, then to justify the number of repeats of Petri dishes in the main erimentah perform correlation analysis seedlings in each petri dish and their set of preliminary experiment, calculating a statistical correlation coefficient.

The method of biotesting for seed germination, for example, red round radish with a white tip when testing river water samples, includes the following steps.

A sampling of river water is carried out, then its conservation and preparation for each test.

In the laboratory, a test is carried out to germinate the seeds of red round radish with a white tip.

To do this, the filter paper is assumed to be round in shape with a diameter of 10 cm Petri dish with a mark on the edge, a template with holes for marking the seed planting places is laid on it, one of the holes being oriented relative to the filter paper mark, with the seed marking being applied with a pencil in the form points.

The template is made in the form of a circle along the diameter of the Petri dish, and the holes for marking the seed seats are placed clockwise after 30 degrees, while the distance between the holes and the center of the template is about 30 mm.

After marking, the template is removed, and the marked filter paper is placed in a Petri dish with the orientation of its label in the north direction along the compass. Do the same with other Petri dishes in 4 repetitions of the experiment. Moreover, before use, Petri dishes are sterilized in an autoclave at 2 atm for 10 minutes or in boiling water for 30 minutes.

Then, in each Petri dish, at the marked places of planting, 12 seeds of red round radish with a white tip are placed on filter paper. Then, 5 ml of the studied water sample is poured into each Petri dish at 4-8-fold repetition, while the liquid level in the cups should be lower than the surface of the seeds. Next, the cup is covered with a lid and placed in a thermostat at a temperature of 20 ° C, and in the absence of a thermostat, an experiment is possible in room conditions, but then, due to temperature fluctuations, it is difficult to compare the results carried out at different times.

After 72 hours of germination of the seeds of red round radish with a white tip before measuring the length of the root for each seedling, the angle of its direction is measured as an indicator of the azimuth of the root, and for non-germinated seeds, the azimuth of the root is taken for a lack of quantitative value and a dash is put in the measurement log.

Subsequently, the length of the root is measured for all seedlings, and this root length is measured for all seeds, including non-germinated seeds, for which the root length is taken to be zero, and the length of the seedlings is measured sequentially, starting from the first seedling clockwise from 30 to 360 degrees.

After completing the test of the test water and taking measurements of the length and azimuth of the roots, the general biotechnical regularity of the distribution of the length of the roots from the azimuth of planting seeds for each Petri dish and their combination is revealed by statistical modeling.

Identification by the method of identification of biotechnological patterns of change in the root length of each plant sprout, depending on the azimuth of planting seedlings for 72 hours of seed germination, is performed according to the generalized formula:

L = L ₁ + L ₂ ,

L ₁ = L ₀ , L ₂ = A cos (πφ / p ± a ₇ ),

, $$p=a_4\pm a_5{\varphi }^{a_6}$$

, $$A=a_{\mathrm{one}}exp(-a_2{\varphi }^{a_3})$$

, $$p=a_{\mathrm{four}}\pm a_5{\varphi }^{a_6}$$

,

where L is the root length of each plant sprout in the Petri dish, mm;

L ₁ - the first component, taking into account the effect of pollution in its pure form, mm;

L ₀ - theoretical independent value of the root length from the azimuth of seed planting, shows that this value is dependent on other factors, mm;

L ₂ - the second component of the vibrational disturbance of the plant along the root length of each seedling in the Petri dish for 72 hours, characterizing the influence of the side of the world, mm;

And - half the amplitude of the vibrational disturbance of the roots of seedlings, mm;

p - half the period of oscillatory changes in azimuth of seed planting, degrees .;

φ is the azimuth of seed planting, deg .;

and ₁ ... a ₇ are the parameters of the finished statistical model, the model parameters take specific values for specific conditions of seed germination with the test water for irrigation.

After identifying the biotechnological pattern of changes in the length of the root of the plant, depending on the azimuth of planting, the significance of each component of the statistical model is determined.

According to the adaptability coefficient, equal to the ratio of the wave second component to the first component of the model, when this coefficient is less than unity, it is estimated that the adaptability of plants to irrigated water is doubtful, and if the unit is exceeded, it is quite reliable.

Then, the correlation coefficient is calculated for all samples of the studied water and for their combination and compared with the control water sample, but at the same time, preliminary and basic experiments are carried out regionally for each water body.

In a preliminary experiment, when comparing with a control sample of pure water, the seeds in the studied contaminated water did not germinate at all or the root length as a percentage of the control was below 70, then the water is not suitable for irrigation, and the prototype threshold is 70% because the soil sorption ability, reduces the inhibitory effect of the studied water, and when the root length in the experiment is more than 120% of the control, it is assumed that the water has stimulating properties, then to justify the number of repeats of Petri dishes in the main The experiments perform a correlation analysis of the seedlings in each Petri dish and from their totality from the preliminary experiment, calculating the statistical correlation coefficient.

Example. The experiments were conducted in February 2009 - November 2010. A water sample was taken from a surface water source in front of the city water intake - the M. Kokshaga river.

Table 1 shows the germination data at room conditions (seeds were planted at 6 p.m. February 3, 2010) in independent samples of 12 seeds of red round radish with a white tip in seven repetitions of Petri dishes.

Table 1 The length of the roots of red round radish in seven Petri dishes, mm Azimuth of landing φ, deg. No. 1 Number 2 Number 3 Number 4 Number 5 Number 6 Number 7 L mm φ _k , deg. L mm φ _k , deg. L mm φ _k , deg. L mm φ _k , deg. L mm φ _to hail. L mm φ _k , deg. L mm φ _k , deg. thirty 43 315 5 225 5 135 26 180 eleven 270 fifteen 315 fifteen 225 60 fifteen 45 eighteen 180 36 135 37 45 41 315 19 225 36 270 90 21 225 8 0 5 315 40 135 25 180 6 45 13 45 120 37 315 25 270 26 315 35 45 eighteen 270 24 90 13 45 150 25 315 9 270 0 - 0 - twenty 135 38 225 eighteen 315 180 fourteen 225 21 90 44 225 0 - 35 180 29th 315 26 225 210 32 315 0 - 17 225 36 45 23 135 27 45 four 315 240 26 315 fifteen 0 0 - 13 225 21 135 41 135 33 135 270 24 315 29th 135 22 315 0 - 17 45 24 135 3 225 300 27 315 38 225 19 315 22 135 5 0 0 - 27 315 330 twenty 45 3 225 0 - 21 135 0 - 19 135 33 45 0 43 45 53 225 22 45 0 - 0 - eleven 0 17 315

During the experiment, a round-shaped filter paper with a mark on the edge fit a template with holes for marking the seed planting places (Fig. 1), which was then placed in a Petri dish with the label pointing northward in the compass. Similarly, all repetitions of Petri dishes were done, after which seeds were placed every 30 degrees of the azimuth of planting φ.

After 72 hours of seed germination before measuring the length of the root, the angle of its direction was measured for each seedling as an indicator of the azimuth of the root, and for the non-germinated seeds, the azimuth of the root was taken for the absence of a quantitative value and a dash was put in the measurement log.

Then, the root length was measured for all seedlings, and this root length was measured for all seeds, including non-germinated seeds, in which the root length value was taken equal to zero.

When watering seeds with river water in one Petri dish (the graph is shown in Fig.6), a two-membered biotechnical regularity was obtained

$$\text{L}={\text{L}}_{\text{one}}+{\text{L}}_{\text{2}}(\mathrm{one})$$

L ₁ = 25.51866,

L ₂ = 35.83023exp (-0.096491φ ^0.51390 ) cos (πφ / (55.91559-0,00050888φ ^1,73960 ) -1,05930).

The first component shows a stable distribution of the length of the roots of plants, that is, is characterized by a stable law (figure 4). The most significant is the second component (figure 5), which characterizes the vibrational disturbance of the red radish seed population from the azimuth of seed planting.

The maximum wave perturbation of seedlings of red round radish 17.54 mm was observed at a seed planting azimuth of 0 degrees.

For the rest of the repetition graphs of the distribution of the length of the root of the radish red round from the bearing azimuth are presented in figures 7-12.

The calculation results by the formula (1) are shown in table 2.

table 2 Change in root length according to a statistical (mathematical) model (1), mm Azimuth of landing φ, deg. Actual Root Length The calculated values of the root length, mm Component Models (1) L ε Δ,% L ₁ BUT R L ₂ thirty 43 42.13 0.87 2.02 25.52 20.59 55.73 16.61 60 fifteen 14.10 0.90 6.00 25.52 16.24 55.28 -11.42 90 21 17.92 3.08 14.67 25.52 13.52 54.64 -7.60 120 37 36.45 0.55 1.49 25.52 11.58 53.81 10.93 150 25 25.42 -0.42 -1.68 25.52 10.09 52.81 -0.10 180 fourteen 17.55 -3.55 -25.36 25.52 8.91 51.65 -7.97 210 32 32.41 -0.41 -1.28 25.52 7.95 50.34 6.90 240 26 23.90 2.10 8.08 25.52 7.14 48.88 -1.62 270 24 23.02 0.98 4.08 25.52 6.46 47.28 -2.50 300 27 29.67 -2.67 -9.89 25.52 5.87 45.54 4.15 330 twenty 21.38 -1.38 -6.90 25.52 5.36 43.67 -4.14 0 43 43.06 -0.06 -0.14 25.52 35.83 55.92 17.54 Note. The maximum relative error is underlined.

The maximum length of the root of the seedlings of red round radish with a white tip is observed with an azimuth of planting of 0 and 30 degrees, a minimum of 180 degrees.

The oscillation amplitude for the best individual of the plant is always zero, that is, the strongest individual is not worried. The large amplitude range, as can be seen from Table 2, shows the expansion of the variability of plant parameters in individual individuals, since river water expands the adaptive possibilities of variability in the growth of red radish roots. At the same time, the amplitude takes on high values when the seeds of the test plant are located in the eastern part of the Petri dish, here the seeds experience great excitement. At 180 ° of the azimuth of planting, the amplitude decreases, reaches a minimum at 330 degrees, while the seeds experience less excitement.

Moreover, half the amplitude takes the least value at 330 degrees, here the plant practically does not worry, it experiences the largest fluctuations on the northern meridian, where the amplitude value is 35.83.

The longer the oscillation period, the easier the adaptation of the population to external conditions occurs due to the fact that the frequency of the vibrational disturbance decreases. Table 2 shows that the frequency of the vibrational disturbance increases from the northern end of the meridian from 0 to 360 degrees.

The maximum relative error in table 2 is underlined and amounts to 14.67%. Therefore, the confidence probability of formula (1) will be no lower than 74.64%. A formula above 70% adequacy is assumed to have a strong relationship between comparable factors. Therefore, formula (1) can be applied for parametric identification from measurement data and from other plants.

The correlation coefficient of 0.9814 is high (Fig.6), which confirms the high adequacy of the obtained statistical model.

Therefore, to describe the distribution of the length of the plant roots from the azimuth of seed planting, a generalized formula is proposed

$$\begin{array}{l}L=L_{\mathrm{one}}+{\mathrm{<figure-callout\; id="2"\; label="L"\; filenames="00000009.png,00000011.png"\; state="\{\{state\}\}">L</figure-callout>}}_2,\\ L_{\mathrm{one}}=L_0,{\mathrm{<figure-callout\; id="2"\; label="L"\; filenames="00000009.png,00000011.png"\; state="\{\{state\}\}">L</figure-callout>}}_2=Acos(\pi \varphi /p\pm a_7),A=a_{\mathrm{one}}exp(-a_2{\varphi }^{a3}),p=a_{\mathrm{four}}\pm a_5{\varphi }^{a6},(2)\end{array}$$

where L is the root length of each plant sprout in the Petri dish, mm;

L ₁ - the first component, taking into account the effect of pollution in its pure form, mm;

L ₀ - theoretical independent value of the root length from the azimuth of seed planting, shows that this value is dependent on other factors, mm;

L ₂ - the second component of the vibrational disturbance of the plant along the root length of each seedling in the Petri dish for 72 hours, characterizing the influence of the side of the world, mm;

A - half the amplitude of the vibrational disturbance of the roots of seedlings, mm;

p - half the period of oscillatory changes in azimuth of seed planting, degrees .;

φ is the azimuth of seed planting, deg .;

a ₁ ... a ₇ - parameters of the finished statistical model, the parameters of the model take specific values for specific conditions of germination of seeds with test water for irrigation.

Comparison of the results of several tests is carried out according to several criteria, quantitatively obtained from experiments and subsequent statistical modeling.

According to table 3, we analyze the limiting values of the individual components and components of the obtained formula (2) according to the results of an experiment conducted in February 2010 in 7 Petri dishes.

Table 3 Significance and adaptability of seedlings in river water Repeat Number Azimuth of landing φ, deg. Component models (2), mm L mm Significance coefficient Adaptability coefficient k L ₂ L ₁ α ₁ α ₂ one thirty 25.52 16.61 43 0.59 0.39 0.6509 60 25.52 -11.42 fifteen 1.70 -0.76 -0.4475 90 25.52 -7.60 21 1.22 -0.36 -0.2978 120 25.52 10.93 37 0.69 0.30 0.4283 150 25.52 -0.10 25 1,02 -0.00 -0.0039 180 25.52 -7.97 fourteen 1.82 -0.57 -0.3123 210 25.52 6.90 32 0.80 0.22 0.2704 240 25.52 -1.62 26 0.98 -0.06 -0.0635 270 25.52 -2.50 24 1.06 -0.10 -0.0980 300 25.52 4.15 27 0.95 0.15 0.1626 330 25.52 -4.14 twenty 1.28 -0.21 -0.1622 0 25.52 17.54 43 0.59 0.41 0.6873 2 thirty 18.22 -1.33 5 3.64 -0.27 -0.0730 60 18.22 -14.64 eighteen 1.01 -0.81 -0.8035 90 18.22 -7.33 8 2.28 -0.92 -0.4023 120 18.22 9.93 25 0.73 0.40 0.5450 150 18.22 12.70 9 2.02 1.41 0.6970 180 18.22 -2.68 21 0.87 -0.13 -0.1471 210 18.22 -13.71 0 - - -0.7525 240 18.22 -4.24 fifteen 1.21 -0.28 -0.2327 270 18.22 11.32 29th 0.63 0.39 0.6213 300 18.22 9.02 38 0.48 0.24 0.4951 330 18.22 -7.42 3 6.07 -2.47 -0.4072 0 18.22 14.32 53 0.34 0.27 0.7859 3 thirty 16.93 -2.01 5 3.39 -0.40 -0.1187 60 16.93 -3.34 36 0.47 -0.09 -0.1973 90 16.93 -2.84 5 3.39 -0.57 -0.1677 120 16.93 -0.77 26 0.65 -0.03 -0.0455 150 16.93 1.72 0 - - 0,1016 180 16.93 3.26 44 0.38 0,07 0.1926 210 16.93 3.01 17 1.00 0.18 0.1778 240 16.93 1.10 0 - - 0.0650 270 16.93 -1.41 22 0.77 -0.06 -0.0833 300 16.93 -3.15 19 0.89 -0.17 -0.1861 330 16.93 -3.15 0 - - -0.1861 0 16.93 0.43 22 0.77 0.02 0,0254 four thirty 18.12 2,57 26 0.70 0.10 0.1418 60 18.12 20.67 37 0.49 0.56 1,1407 90 18.12 24.19 40 0.45 0.60 1.3350 120 18.12 7.02 35 0.52 0.20 0.3874 150 18.12 -18.22 0 - - -1.0055 180 18.12 -18.38 0 - - -1.0143 210 18.12 15.60 36 0.50 0.43 0.8609 240 18.12 6.22 13 1.39 0.48 0.3433 270 18.12 -13.07 0 - - -0.7213 300 18.12 -2.15 22 0.82 -0.10 -0.1187 330 18.12 5.51 21 0.86 0.26 0.3041 0 18.12 -17.36 0 - - -0.9581 5 thirty 19.28 -3.79 eleven 1.75 -0.34 -0.1966 60 19.28 12.51 41 0.47 0.31 0.6489 90 19.28 6.42 25 0.77 0.26 0.3330 120 19.28 -5.40 eighteen 1,07 -0.30 -0.2801 150 19.28 -5.65 twenty 0.96 -0.28 -0.2930 180 19.28 1.38 35 0.55 0.04 0,0716 210 19.28 3.85 23 0.84 0.17 0.1997 240 19.28 0.45 21 0.92 0.02 0.0233 270 19.28 -2.17 17 1.13 -0.13 -0.1126 300 19.28 -0.99 5 3.86 -0.20 -0.0513 330 19.28 0.98 0 - - 0,0508 0 19.28 -22.90 0 - - -1.1878 6 thirty 15.23 -2.11 fifteen 1,02 -0.14 -0.1385 60 15.23 0.37 19 0.80 0.02 0,0243 90 15.23 3.40 6 2.54 0.57 0.2232 120 15.23 7.04 24 0.63 0.29 0.4622 150 15.23 11.29 38 0.40 0.30 0.7413 180 15.23 15.87 29th 0.53 0.55 1,0420 210 15.23 19.80 27 0.56 0.73 1,3001 240 15.23 20.17 41 0.37 0.49 1,3244 270 15.23 10.21 24 0.63 0.43 0.6704 300 15.23 -15.40 0 - - -1.0112 330 15.23 3.78 19 0.80 0.20 0.2482 0 15.23 -4.15 eleven 1.38 -0.38 -0.2725 7 thirty 19.19 -0.91 fifteen 1.28 -0.06 -0.0474 60 19.19 5.04 36 0.53 0.14 0.2626 90 19.19 0.79 13 1.48 0.06 0.0412 120 19.19 -8.00 13 1.48 -0.62 -0.4169 150 19.19 6.14 eighteen 1,07 0.34 0.3200 180 19.19 4.14 26 0.74 0.16 0.2157 210 19.19 -14.69 four 4.80 -3.67 -0.7655 240 19.19 18.59 33 0.58 0.56 0.9687 270 19.19 -14.06 3 6.40 -4.69 -0.7327 300 19.19 3.20 27 0.71 0.12 0.1668 330 19.19 10.84 33 0.58 0.33 0.5649 0 19.19 -3.32 17 1.13 -0.20 -0.1730

The adaptability of the red radish seed population (Table 3) to external influences is shown by the coefficient k. Moreover, adaptability can be crisis-negative or crisis-positive. Under the condition k> 1, we will consider the adaptability of plants to irrigated water doubtful, and under the condition k <1, it is sufficiently reliable.

From the data in Table 3, it can be seen that the greatest adaptability of 1.3350 in the direction of positive adaptation is that the seeds of the red round radish have an azimuth of planting of 90 degrees. The adaptability coefficient, calculated by the formula k = L _max2 / L _max1 , is equal to _1.3350 with this arrangement of the seed. The smallest positive effect of 0.0039 is observed at 150 °.

The maximum crisis for seedlings is observed at 0 degrees with negative adaptation to growth conditions with an adaptability coefficient of k = -1.1878, that is, when the seeds of red radish were located in the northern part of the Petri dish. However, it is less than the positive high of 1.3350.

To justify the number of repeats of Petri dishes in the main experiments, we perform a correlation analysis of the seedlings in each Petri dish according to well-known standard methods.

Table 4 presents the number of seeds sprouted according to the results of the experiment on germination of red round radish seeds with a white tip for four months.

Table 4 The number of seeds sprouted from the azimuth of their planting for four months No. Azimuth of landing φ, deg. The number of seeds sprouted 12/02/09 04/05/10 08/10/10 10/13/10 one thirty 7 7 7 7 2 60 5 7 four 7 3 90 7 5 6 6 four 120 5 6 5 6 5 150 7 7 5 7 6 180 6 7 5 7 7 210 6 6 four 7 8 240 6 6 6 6 9 270 6 7 2 6 10 300 7 5 four 7 eleven 330 6 7 6 6 12 0 6 7 6 7

We will calculate the indicators of descriptive statistics for the subsequent justification of the required sample size.

The results of statistical processing are presented in table 5.

The coefficient of variation V _σ was less than 33%, that is, all the aggregates are homogeneous, the fluctuations in the length of the root of the red round radish are insignificant.

According to the results obtained, E <0, that is, the data is more evenly distributed over the entire range of values from x _max to x _min .

Moreover, the distribution is flat.

For August, E> 0, the data are concentrated around the arithmetic mean value. Since As <0, left-side asymmetry is observed.

The essence of the method for calculating the required number of observations is that sets of factor values are taken as statistical samples. For any number of factors, the ratio

n = m

where n is the total number of seeds in the Petri dish for all repetitions of the experiment,

m is the number of seeds sprouted in all repetitions of the Petri dishes,

m 'is the number of repeats of Petri dishes.

Table 5 Descriptive statistics on the number of seeds sprouted red round red radish by month Statistical indicators Monthly Descriptive Statistics Values 12/02/09 04/05/10 08/10/10 10/13/10 Swing R, mm 2.00 2.00 5.00 1.00 Minimum x _min , mm 5.00 5.00 2.00 6.00 Maximum x _max , mm 7.00 7.00 7.00 7.00

, ммAverage $$\overline{x}$$

mm 6.17 6.42 5.00 6.58 Average sampling error, mm 0.21 0.23 0.39 0.15 Standard deviation σ, mm 0.72 0.79 1.35 0.51 Sample variance σ ² 0.52 0.63 1.82 0.27 Coefficient of variation V _σ ,% 11.64 12.36 29.97 7.82 Excess E -0.69 -0.46 0.92 -2.26 Asymmetry As -0.26 -0.99 -0.80 -0.39 Limit sampling error at P = 0.954 Δ _x 0.46 0.50 0.86 0.33

To calculate the interval Δx, we apply the formula G.A. Strages

$$\Delta x=\frac{x_{max}-x_{min}}{\mathrm{one}+3.322\mathrm{one}gn},(3)$$

where x _max , x _min - the maximum and minimum number of seeds sprouted.

The expression in the denominator (3) characterizes the number of observation groups m. To calculate the number of observations, we use the formula

$$n\ge l\cdot [m\text{'}\text{'}],(\mathrm{four})$$

where [m '] is the number of repetitions of Petri dishes in our experiment. For example, the current number of observations n _m was 84. Then the number of observation groups is l = 7. We conducted an experiment with seven repetitions, so we take [m '] equal to seven. Then we get n≥7 × 7 = 49. Since n _m > n, then the initial data are sufficient, the condition n≥l · [m '] is satisfied for this factor. Let n _m = 48; therefore, l = 7. The number of repetitions will be equal to four. Since n _m = 7 × 4 = 28, the condition n≥l · [m '] is satisfied.

It follows that the main experiments on biotesting river water in the city of Yoshkar-Ola are enough to carry out when germinating seeds of red round radish with a white tip in an amount of 12 pieces in four repetitions in Petri dishes.

Thus, the method of biotesting for seed germination for river water in the city of Yoshkar-Ola main experiments on germinating red round radish seeds with a white tip include uniform seed placement in a Petri dish in the amount of 12 pieces in place after 30 degrees of planting azimuth with four repeats of Petri dishes .

The proposed method describes with high accuracy the influence of azimuth on the distribution of the root length of the seedling when determining the pollution of river water. It has the simplicity of testing river water, as well as filtration water, which significantly increases the accuracy of correlating pollution measurements in rivers and reservoirs with the results of water quality testing by plants. Therefore, the proposed method allows for some time to abandon the costly methods of analyzing water samples and replace them with the proposed methods for a comprehensive assessment of water quality.

The application of the proposed method expands the possibilities of territorial environmental pollution monitoring by simple means at those waterways of the river network where settlements and agricultural land plots, landfills and solid waste storage facilities are located.

Claims (4)

1. A method of placing seeds in a Petri dish during biotesting of river water, including uniformly placing seeds of red round radish with a white tip on filter paper in a Petri dish, into which 5 ml of the studied water sample is poured, while the liquid level in the cup should be lower than the surface seed, characterized in that the filter paper is made round in diameter by the diameter of the Petri dish with a mark on the edge, a template with holes for marking the places for planting seeds is laid on it, one of the holes being oriented Filter paper marks, then marked filter paper is placed in a Petri dish with the compass pointing northward along the compass, then seeds are placed on the marked Petri dish on marked places of planting, the test water is poured, after germination of the seeds, before seedlings are taken from the Petri dish To measure the root length of each seedling, the angle of the direction of its root is measured using a compass as an indicator of the azimuth of the root, moreover, the root azimuth of non-sprouted seeds is taken for the absence of quantitative This value is also marked with a dash in the measurement log; later, the root length is measured for all seedlings in a Petri dish, and this length is taken to be zero for non-sprouted seeds. 2. The method of placing seeds in a Petri dish during biotesting of river water according to claim 1, characterized in that a template with holes for marking the places for planting seeds is laid on a round filter paper with the diameter of the Petri dish with a mark on the edge, and one of the holes is oriented relatively labels of filter paper, while the marks of the places of planting seeds are applied in pencil in the form of dots, after drawing the marking, the template is removed, the marked filter paper is placed in a Petri dish with the orientation of its label in the north direction mpasu. 3. The method of placing seeds in a Petri dish during biotesting of river water according to claim 1, characterized in that the template for planting seeds is made in the form of a circle in diameter of the Petri dish, for example, from a transparent thin sheet of plastic, with holes for marking the places for planting seeds clockwise through 30 ° with writing numbers near the holes, while for Petri dishes with a diameter of 100 mm, the distance between the centers of the holes and the center of the template is taken to be approximately 30 mm, and the diameters of the holes are large compared to the diameters and seeds of red round radish with a white tip. 4. The method of placing seeds in a Petri dish during biotesting of river water according to claim 1, characterized in that the root length is measured for all seedlings, and this root length is measured for all seeds, including non-germinated seeds, in which the root length value is assumed to be zero moreover, the measurement of the length of the seedlings is carried out sequentially clockwise, starting from the first seedling in a clockwise direction from 30 to 360 °.

Images