CN110352850B - Method for breeding high-yield, high-quality and stress-resistant excellent new species of common andrographis herb - Google Patents
Method for breeding high-yield, high-quality and stress-resistant excellent new species of common andrographis herb Download PDFInfo
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Abstract
The invention discloses a method for breeding a new variety of andrographis paniculata with high yield, high quality, stress resistance and excellent quality. The method comprises the following steps: the seed of common andrographis herb is used as the material,60the Co-gamma rays are used as radiation sources to be radiated and then are divided into 3 groups,respectively soaking seeds in distilled water, germinating in salt stress environment containing NaCl solution and drought stress environment containing PEG6000 solution, and calculating germination rate, germination vigor, germination index, activity index, root length, bud length or fresh weight; and screening the high-yield high-quality stress-resistant excellent new variety with salt resistance and drought resistance according to the change of the germination rate, the germination vigor, the germination index, the vitality index, the root length, the bud length and the fresh weight of each sample. The invention shows that the proper dosage is adopted60Co-gamma ray irradiation of the andrographis paniculata seeds is expected to become a good method for respectively improving tolerance of the andrographis paniculata under salt stress and drought stress in practice, and has great application potential; the invention has high application value for relieving the damage of the stress to plants.
Description
Technical Field
The invention belongs to the technical field of biological planting. More particularly relates to a method for breeding a new variety of andrographis paniculata with high yield, high quality, stress resistance and excellent quality.
Background
The Andrographis paniculata is derived from Andrographis paniculata (Burm.f.) Nees of Acanthaceae, and is applied to the medicine from the aerial part, and due to the serious phenomena of variety degradation, yield reduction, content reduction of effective components and the like caused by long-term artificial cultivation, the research of cultivating new germplasm with high yield, high effective components and high quality is urgent. The radiation mutagenesis is mainly to make the material produce mutation by radiating the material with a certain dose of ray, induce mutation, and finally breed a new variety breeding process which is valuable for scientific research or production from the variant material through screening, measuring and selecting. The irradiation mutagenesis technology has the advantages of high mutation rate, wide mutation spectrum, stable progeny character, short breeding period and the like, and is one of effective ways for obtaining new germplasm resources at present. Gamma rays are one of the radiation mutagenic rays and the radiation source is60Co, a neutral ray, has strong penetration ability, relatively uniform radiation dose and can treat a large amount of materials at one time.60Co-gamma radiation is the most commonly used irradiation radiation, with mutagenesisLow cost, high mutation rate, strong penetrating power, easy to cause plant morphological structure and physiological and biochemical variation, etc. and has wide breeding foreground.
At present, tissue culture seedlings of ornamental plants such as chrysanthemum, hosta plantaginea, Chinese rose and the like are developed60Co-gamma ray radiation mutation breeding method and related research of semilethal dose, such as influence of radiation on growth of chrysanthemum tissue culture seedling [ J ] when the semilethal dose of chrysanthemum tissue culture seedling is 20Gy (Li, Hujinping. radiation]Research on homeland and natural resources, 2011); semi-lethal dose of 5Gy (Li, quyangting, Chenfei, Hosta ventricosa tissue culture seedling) [ J ] radiation breeding research]Forestry science 2014); the semi-lethal dose of the tissue culture seedling of the rose is 50Gy (Zhang Xing, Tang Huan Wei, che Dynasty. the rose60Co-gamma radiation breeding research and preliminary analysis of progeny variation [ J]Research of homeland and natural resources, 2010); chinese patent document CN 103975861A discloses fern green spheroids60The Co-gamma ray radiation mutagenesis method takes green spheroids generated by the induction of aseptic seedlings of ferns as materials under the condition of isolated culture,60co-gamma ray is used as radiation source, provides new technology for performing radiation mutation breeding of ferns on organ level, and is suitable for breeding new fern variety.
However, radiation mutation breeding is a complex process involving stages of different phenotypes, each stage being dependent on both bacterial genes and host plant genes. At present, related researches on influences of biological nitrogen fixation and nodulation nitrogen fixation of non-leguminous medicinal materials on the quality of medicinal materials are few, the influence researches on the andrographis paniculata are rare, and the development and utilization of rhizobium resources and the improvement of the quality of andrographis paniculata raw medicinal materials are urgent market requests.
Radiation mutagenesis of modern chinese herbal plants also presents several problems. Firstly, the genetic background of the Chinese herbal medicine plants is easily and seriously damaged by irradiation mutagenesis; secondly, it is difficult to determine the optimal mutagen and the mutagenic dose required by different plants in the process of radiation mutation breeding. Thus, the selection of radiation mutagens and mutagenic doses is much more blind and can only be determined by repeated screening through experimentation.
Disclosure of Invention
The technical problem to be solved by the invention is to overcome the problems of degeneration of andrographis paniculata varieties and reduction of yield and quality in the prior art, and provide a method for breeding high-yield high-quality stress-resistant excellent new varieties of andrographis paniculata.
Another objective of the invention is to provide an application of the method in breeding salt-tolerant and/or drought-tolerant andrographis paniculata.
The above purpose of the invention is realized by the following technical scheme:
a method for breeding a new variety of andrographis paniculata with high yield, high quality, stress resistance and excellent quality comprises the following steps:
s1, material preparation: the seed of common andrographis herb is used as the material,60co-gamma ray is used as radiation source to make radiation treatment,60the irradiation dose of the Co-gamma ray is 10-300 Gy;
s2, dividing the andrographis paniculata seeds subjected to radiation mutagenesis treatment into 3 groups, carrying out distilled water seed soaking treatment on a control group for germination, carrying out germination on a first test group in a salt stress environment containing a NaCl solution, carrying out germination on a second test group in a drought stress environment containing a PEG6000 solution, calculating the germination vigor after 3-4 days of germination, calculating the germination rate after 7-8 days of germination, and calculating the germination index, the vitality index, the root length, the bud length or the fresh weight;
s3, screening a new high-yield high-quality stress-resistant excellent variety with salt resistance and drought resistance according to the change of the germination rate, the germination vigor, the germination index, the vitality index, the root length, the bud length and the fresh weight of each sample of the salt stress treated seed and the drought stress treated seed.
Preferably, the first and second electrodes are formed of a metal,60the irradiation dose of the Co-gamma ray is 50-100 Gy. The experimental result shows that the content of andrographolide in the 50Gy irradiation dose group is the highest and is 2.7514%, which is 175.14% of that in the control group; the yield per mu of the common andrographis herb in the 100Gy irradiation dose group is highest and is 128.147Kg, which is 143.08% of that in the control group; the salt tolerance and drought resistance of the andrographis paniculata 50Gy irradiation dose group are optimal.
Preferably, the germination is performed in a salt stress environment as follows: soaking the andrographis paniculata seeds subjected to radiation mutagenesis treatment in treatment solutions respectively: distilled water, 0.1% NaCl, 0.2% NaCl, 0.3% NaCl, 0.4% NaCl, 0.5% NaCl, 0.6% NaCl, 0.7% NaCl, 0.8% NaCl, 0.9% NaCl, 1.0% NaCl solution for 24-30 h.
Preferably, the germination in drought stress environment is performed as follows: soaking the andrographis paniculata seeds subjected to radiation mutagenesis treatment in treatment solutions respectively: distilled water, 10% PEG6000, 12% PEG6000, 15% PEG6000, 18% PEG6000 and 21% PEG6000 solution for 2-4 h.
Preferably, after soaking the obtained salt-stress and drought-stress andrographis paniculata seeds, absorbing the attached water by using sterile filter paper, respectively placing the seeds into culture dishes paved with double layers of filter paper, and placing the seeds into an illumination incubator to perform germination culture experiments; wherein salt stress 5mL of salt solution of corresponding concentration was added to each dish.
Preferably, the conditions of the germination culture are: the temperature is 25 +/-2 ℃, the illumination intensity is 10000-12000 lx, and the illumination time is 12h light/12 h dark.
Preferably, distilled water is supplemented every 1d during the germination of the salt-stressed seeds, so that the salt concentration is kept constant in the germination process of the seeds; the PEG6000 solution is replaced every 2d during the germination period of the drought stress seeds, so that the water potential is kept constant in the seed germination process.
Preferably, 20-50 seeds are cultured in each culture dish, and the experiment is repeated for 3-5 times.
Preferably, in60Before Co-gamma ray radiation, pretreatment is carried out on the andrographis paniculata seeds: selecting seeds which are not damaged and have consistent sizes, disinfecting the seeds for 20-25 min by 0.5-1% of sodium hypochlorite respectively, washing the seeds for 3-5 times by distilled water, and sucking the attached water by filter paper.
The application of the method in the breeding of the salt-tolerant and/or drought-tolerant andrographis paniculata also falls within the protection scope of the invention.
Compared with the prior art, the invention has the following beneficial effects:
the invention shows that the proper dosage is adopted60Co-gamma ray irradiation of the andrographis paniculata seeds is expected to become a good method for respectively improving tolerance of the andrographis paniculata under salt stress and drought stress in practice, and has great application potential. The invention can breed the new germplasm of the common andrographis herb with high yield, high quality and excellent stress resistance, and is richThe method has reasonable design, reliable result and popularization value, is one of effective ways for solving various problems of the common andrographis herb, such as variety degradation, yield and quality reduction and the like, and provides theoretical basis and technical basis for enriching the common andrographis herb germplasm resources.
Drawings
FIG. 1 is a high performance liquid phase diagram of andrographolide content in a 50Gy radiation dose group.
FIG. 2 is a high performance liquid phase diagram of dehydroandrographolide content in a 50Gy radiation dose group.
FIG. 3 is the effect of different NaCl concentrations on root length and shoot length of a seedling of Andrographis paniculata; wherein, 1-3: control group Andrographis paniculata seedlings; 4-6: andrographis paniculata seedlings stressed with 0.1% NaCl; 7-9: andrographis paniculata seedlings stressed with 0.2% NaCl; 10-13: andrographis paniculata seedlings stressed with 0.3% NaCl.
Detailed Description
The invention is further described with reference to the drawings and the following detailed description, which are not intended to limit the invention in any way. Reagents, methods and apparatus used in the present invention are conventional in the art unless otherwise indicated.
Unless otherwise indicated, reagents and materials used in the following examples are commercially available.
Example 160Influence of Co-gamma ray on yield per mu of andrographis paniculata
1. Method of producing a composite material
Selecting full and uniform herba Andrographitis seed, cleaning with deionized water, removing suspended shriveled granule and impurity, sterilizing with 1% sodium hypochlorite for 20min, washing with distilled water for 3 times, sucking off attached water with filter paper, and adding different dosages60After Co-gamma ray irradiation (0, 10, 20, 50, 100, 200, 300Gy), 100 granules of the andrographis paniculata seeds of each irradiation dose group are taken and soaked in distilled water for 24 hours. Sowing the common andrographis herb seeds of each dosage group into 50-hole seedling pots, wherein each 1 seed is placed in one hole, and each group comprises two seedling pots. Supplementing water in seedling-raising pot at 12 am and 5 pm every day to keep the culture soil moist, observing herba AndrographitisAnd (3) the growth condition of the seedlings, namely randomly selecting 20 andrographis paniculata seedlings to be transplanted into a greenhouse when a third pair of true leaves grow. Piercing plants were harvested at the beginning of flowering, and the fresh and dry weights of individual plants of Andrographis paniculata Nees were determined using an analytical balance, the results are shown in Table 1.
2. Results
TABLE 1 statistical analysis of the fresh weight of andrographis paniculata plants at different irradiation doses (n ═ 3)
p is less than 0.05, and the difference is significant.
As can be seen from Table 1, p is less than 0.05, i.e., different irradiation doses have significant influence on the fresh weight of Andrographis paniculata plants, and the fresh weight of Andrographis paniculata plants tends to increase first and then decrease with increasing irradiation dose. Wherein the fresh weight of the andrographis paniculata plant in the 100GY dosage group is about 143.08% of that in the control group; 50GY dose group; and the minimum of the 300GY dose group was about 46.61% of the control group. Therefore, 50-100 Gy is the optimal irradiation dose for breeding the andrographis paniculata with high yield and quality.
Example 260Influence of Co-gamma ray on the content of effective components of andrographis paniculata
1. Method of producing a composite material
Selecting andrographis paniculata plants with good growth conditions, and measuring the content of active ingredients by using a drug dictionary method: octadecylsilane chemically bonded silica is used as a filling agent; methanol-water (52: 48) is used as a mobile phase; the detection wavelength of andrographolide is 225nm, and the detection wavelength of dehydroandrographolide is 254 nm. The number of theoretical plates is not less than 2000 calculated according to the peak of andrographolide and dehydroandrographolide. Accurately weighing appropriate amount of andrographolide control substance and dehydroandrographolide control substance, and adding methanol to obtain mixed solution containing 0.1mg of andrographolide per mL. Taking about 0.5g of the powder of the andrographis paniculata plant sample subjected to radiation mutagenesis treatment in example 1 (screened by a No. four sieve), precisely weighing, placing in a conical flask with a plug, precisely adding 25mL of 40% methanol, weighing, soaking for 1h, ultrasonically treating (power 250W, frequency 33kHz) for 30min, cooling, weighing again, supplementing the lost weight with 40% methanol, shaking uniformly, and filtering. Precisely measuring 10mL of the subsequent filtrate, eluting with 15mL of methanol on a neutral alumina column (200-300 meshes, 5g, inner diameter of 1.5cm), collecting the eluent, putting the eluent in a 50mL measuring flask, adding methanol to the scale, and shaking up to obtain the final product. Then precisely sucking 5 μ L of each of the reference solution and the sample solution, injecting into a liquid chromatograph, and measuring. The results are shown in Table 2.
2. Results
FIG. 1 is a high performance liquid phase diagram of andrographolide content in a 50Gy radiation dose group. FIG. 2 is a high performance liquid phase diagram of dehydroandrographolide content in a 50Gy radiation dose group. A large number of preliminary studies show that different irradiation doses have great influence on the determination result of the content of the effective components of the andrographis paniculata.
TABLE 2 mu yield of the effective components of the aerial parts of Andrographis paniculata Nees in different dosages (n is 3)
p is less than 0.05, and the difference is significant.
As can be seen from Table 2, p is less than 0.05, i.e., different irradiation doses have significant influence on the content of effective components of Andrographis paniculata Nees, wherein the content of andrographolide in the 50Gy irradiation dose group is 2.7514% which is 175.14% of that in the control group; 100Gy irradiation dose group; therefore, aiming at the breeding of the high-quality new variety of the common andrographis herb, the optimal irradiation dose is 50-100 Gy.
Example 3 Andrographis paniculata salt tolerance test
1. Salt stress test
(1) Method of producing a composite material
The salt was set to (0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0%) NaCl solution (g/mL). Soaking herba Andrographitis seeds in NaCl solutions with different concentrations for 24 hr, taking out, and drying with filter paper. Uniformly placing the soaked seeds into culture dishes paved with 2 layers of filter paper, and adding 5mL of NaCl solution with corresponding concentration (the concentration of the salt solution is consistent with that of the soaked seeds) into each culture dish; and (3) placing the culture dish in an illumination incubator for culture, wherein the temperature is 25 ℃, the illumination intensity is 10000-12000 lx, and the illumination time is 12h light/12 h dark. 50 seeds were treated in each treatment group, 3 replicates were provided, and distilled water was added once a day to ensure that the respective salt concentration remained constant during seed germination. The germination condition of the seeds is counted every 24h (taking the radicle protruding the seed coat by 1mm as a standard), and the counting is carried out for 7 days. And counting growth indexes such as germination vigor, germination indexes, vitality indexes, root length, bud length, new weight and the like, wherein the experimental results are shown in a table 3.
(2) Results
TABLE 3 statistical analysis of various growth indices of the seeds of Andrographis paniculata with different NaCl concentrations (n ═ 3)
p is less than 0.05, and the difference is significant.
As can be seen from Table 3, p is less than 0.05, i.e., different NaCl concentrations have significant influence on the growth and development of the andrographis paniculata. Under the stress of NaCl with different concentrations, the growth condition of the andrographis paniculata seeds is influenced by different degrees, and each growth index of the andrographis paniculata seeds is in a descending trend along with the increase of the NaCl concentration. Compared with a control group, the difference of the growth indexes of the germination potential, the root length and the fresh weight is smaller when the NaCl with the concentration of 0.1% is treated; under the treatment of 0.2% NaCl, the growth indexes of germination vigor, germination index, vitality index, root length, bud length, fresh weight and the like are remarkably different from those of a control group, and are respectively 48.39, 63.04, 35.90, 12.31, 43.75 and 56.52% of the control group; under the treatment of 0.3% and 0.4% NaCl, the growth indexes of germination vigor, germination index, vigor index, root length, bud length, fresh weight and the like are obviously different from those of a control group, but most of the andrographis paniculata seeds cannot grow into normal seedlings, and under the treatment of NaCl salt solution with the concentration of more than 0.4%, the germination rate of the andrographis paniculata seeds is 0, which indicates that the salt stress degree is too heavy. In conclusion, 0.2% NaCl can be used as a suitable treatment concentration for identifying the salt tolerance of the Andrographis paniculata Nees seeds.
2、60Effect of Co-gamma ray on salt-stressed Andrographis paniculata
(1) Method of producing a composite material
The andrographis paniculata seeds of the group of different radiation doses in example 1 are soaked for 24h with NaCl solution with appropriate concentration respectively. The soaked seeds are uniformly placed into culture dishes paved with 2 layers of filter paper, 5mL of NaCl solution with corresponding concentration (the concentration of the salt solution is consistent with that of the soaked seeds) is added into each culture dish, and the culture dishes are placed into a light incubator for culture. 50 seeds were treated in each treatment group, 3 replicates were provided, and distilled water was added once a day to ensure that the respective salt concentration remained constant during seed germination. The germination condition of the seeds is counted every 24h (taking the radicle protruding the seed coat by 1mm as a standard), the counting is carried out for 7 days, and the experimental results are shown in a table 4.
(2) Results
TABLE statistical analysis of the growth indices of Andrographis paniculata seeds in different dose groups at 40.2% NaCl concentration (n-3)
p is less than 0.05, and the difference is significant.
FIG. 3 shows the effect of different NaCl concentrations on the bud length and root length of Andrographis paniculata. As can be seen from FIG. 3 and Table 4, the growth status of Andrographis paniculata Nees seeds of different irradiation dose groups is affected to different extent under the stress of 0.2% NaCl, and the fresh weight, root length and bud length of Andrographis paniculata Nees of 50GY dose group are the largest and respectively 137.50%, 129.69% and 102.78% of the control group, and are the second best irradiation dose under the stress of 0.2% NaCl salt and 100GY dose group.
Example 4 Andrographis paniculata drought resistance test
1. Drought stress experiment
Setting PEG concentrations of 10%, 12%, 15%, 18% and 21% to 5 treatment groups, using distilled water for blank groups, and investigating the influence of different concentrations of PEG6000 on the drought stress of the andrographis paniculata seeds. Selecting 80 pretreated seeds, dividing the seeds into 4 groups, immersing the seeds into 20 groups of treatment solutions, soaking the seeds for 2 hours at room temperature, taking the seeds out, sucking the water on the surface by using filter paper, inoculating the seeds into a culture dish paved with 2 layers of filter paper to perform germination culture experiments, repeating the steps for 3 times for each treatment group, observing, recording and calculating the growth indexes of germination vigor, germination index, vitality index, root length, bud length, new weight and the like of the seeds of each group, wherein the experimental results are shown in a table 5.
(2) Results
TABLE 5 statistical analysis of various growth indices of the seeds of Andrographis paniculata at different PEG6000 concentrations (n ═ 3)
p is less than 0.05, and the difference is significant.
As can be seen from Table 5, the growth status of Andrographis paniculata Nees seeds is significantly affected under PEG stress at different concentrations, and the growth indexes of Andrographis paniculata Nees seeds decrease with increasing concentration of PEG 6000. The difference of the germination rates of the 10%, 12% and 15% treatments is smaller than that of the control group, and the germination rates of the 18% and 21% treatment groups are obviously reduced; with the increase of PEG concentration, the growth indexes of the andrographis paniculata seeds, such as germination vigor, root length and the like, are in a descending trend. The germination potential and the root length of the andrographis paniculata in the 12% treatment group are obviously different from those in the control group, and are respectively 64.90% and 49.86% of those in the control group; under the PFG treatment of more than 12 percent, the growth indexes of germination vigor, root length, fresh weight and the like are obviously different from those of a control group, but most of the andrographis paniculata seeds can not grow into normal seedlings, which indicates that the drought stress degree is too heavy. In conclusion, 12% PEG can be used as a suitable treatment concentration for identifying drought tolerance of Andrographis paniculata Nees seeds.
2、60Effect of Co-gamma ray on drought stress Andrographis paniculata
Placing the common andrographis herb seeds of different radiation dose groups in a 9cm culture dish padded with 2 layers of filter paper, respectively adding 5mL of PEG6000 solution with proper concentration into the culture dish, and then placing the culture dish in a constant-temperature incubator at 25 ℃ for culture, wherein the illumination condition is 12h light/12 h dark, the illumination intensity is 10000-12000 lx, 3 times of each treatment group are repeated, and 50 seeds are repeated. Distilled water treated seeds were used as controls. The germination bed treated with PEG6000 solution was changed every 2d during germination to keep the water potential constant. The germination is measured when the radicle breaks through 1mm of the seed coat, and the initial stage of germination is taken as the day when the earliest seed in 3 times of repetition germinates. And counting growth indexes such as germination vigor, germination indexes, vitality indexes, root length, bud length, new weight and the like, wherein the experimental results are shown in a table 6.
(1) Method of producing a composite material
Placing the common andrographis herb seeds of different radiation dose groups in example 1 in a 9cm culture dish padded with 2 layers of filter paper, respectively adding 5mL of PEG6000 solution with proper concentration in the culture dish, and then placing the culture dish in a constant temperature incubator for culture, wherein the illumination condition is 12h light/12 h dark, the illumination intensity is 10000-12000 lx, and each treatment group has 3 repetitions, and each repetition has 50 seeds. Distilled water treated seeds were used as controls. The germination bed treated with PEG6000 solution was changed every 2d during germination to keep the water potential constant. The germination is measured when the radicle breaks through 1mm of the seed coat, and the initial stage of germination is taken as the day when the earliest seed in 3 times of repetition germinates. And counting growth indexes such as germination vigor, germination indexes, vitality indexes, root length, bud length, new weight and the like, wherein the experimental results are shown in a table 6.
(2) Results
TABLE 612 statistical analysis of the growth indices of Andrographis paniculata Nees in different dose groups at% PEG6000 concentration (n-3)
p is less than 0.05, and the difference is significant.
As can be seen from Table 6, the growth conditions of Andrographis paniculata Nees in different dose groups were affected to different degrees under 12% PEG6000 stress, and the fresh weight and root length of Andrographis paniculata Nees in 50GY dose group were the highest, 106.90% and 103.92% of the control group, respectively, and the optimal irradiation dose under 12% PEG6000 drought stress.
As can be seen from the above examples, suitable dosages are employed60Co-gamma ray irradiation of andrographis paniculata seeds can effectively promote germination of andrographis paniculata seeds and growth of seedlings under different adversity stresses, increase accumulation of biomass of andrographis paniculata seedlings, respectively improve germination rate, germination vigor, germination index, vitality index, root length, bud length and fresh weight of andrographis paniculata under salt and drought stresses, and relieve salt stress and drought stress on seedsAnd damage to the plant. By using60The Co-gamma ray irradiation of the andrographis paniculata seeds is easy to operate and effective in practice, is expected to become a good method for effectively improving the germination rate of the andrographis paniculata seeds and respectively improving the tolerance of the andrographis paniculata seeds under salt stress and drought stress in practice, and has application potential
Is very large.
The above detailed description is of the preferred embodiment for the convenience of understanding the present invention, but the present invention is not limited to the above embodiment, that is, it is not intended that the present invention necessarily depends on the above embodiment for implementation. It will be apparent to those skilled in the art that any modification of the present invention, equivalent substitutions of selected materials and additions of auxiliary components, selection of specific modes and the like, which are within the scope and disclosure of the present invention, are contemplated by the present invention.
Claims (7)
1. A method for breeding a new high-yield high-quality stress-resistant excellent common andrographis herb variety is characterized by comprising the following steps:
s1, material preparation: taking andrographis paniculata seeds as a material, taking 60 Co-gamma rays as a radiation source for radiation treatment, wherein the radiation dose of the 60 Co-gamma rays is 50-100 Gy;
s2, dividing the andrographis paniculata seeds subjected to radiation mutagenesis treatment into 3 groups, carrying out distilled water seed soaking treatment on a control group for germination, carrying out germination on a first test group in a salt stress environment containing a NaCl solution, carrying out germination on a second test group in a drought stress environment containing a PEG6000 solution, calculating the germination vigor after 3-4 days of germination, calculating the germination rate after 7-8 days of germination, and calculating the germination index, the vitality index, the root length, the bud length or the fresh weight;
s3, screening high-yield high-quality stress-resistant excellent new varieties with salt resistance and drought resistance according to the change of the germination rate, the germination vigor, the germination index, the vitality index, the root length, the bud length and the fresh weight of each sample of the salt stress treated seeds and the drought stress treated seeds;
the germination under the salt stress environment is as follows: soaking the andrographis paniculata seeds subjected to radiation mutagenesis treatment in treatment solutions respectively: distilled water, 0.1% NaCl, 0.2% NaCl, 0.3% NaCl, 0.4% NaCl, 0.5% NaCl, 0.6% NaCl, 0.7% NaCl, 0.8% NaCl, 0.9% NaCl, 1.0% NaCl solution for 24-30 h;
the germination under the drought stress environment comprises the following steps: soaking the andrographis paniculata seeds subjected to radiation mutagenesis treatment in treatment solutions respectively: distilled water, 10% PEG6000, 12% PEG6000, 15% PEG6000, 18% PEG6000 and 21% PEG6000 solution for 2-4 h.
2. The method of claim 1, wherein after soaking the obtained salt-stress and drought-stress andrographis paniculata seeds, the seeds are dried by using sterile filter paper, attached with water, placed in culture dishes paved with double layers of filter paper, and placed in an illumination incubator for germination culture experiments; wherein salt stress 5mL of salt solution of corresponding concentration was added to each dish.
3. The method of claim 2, wherein the conditions of the germination culture are: the temperature is 25 +/-2 ℃, the illumination intensity is 10000-12000 lx, and the illumination time is 12h light/12 h dark.
4. The method according to claim 2, wherein distilled water is supplemented every 1d during germination of the salt-stressed seeds to keep the salt concentration constant during germination of the seeds; the PEG6000 solution is replaced every 2d during the germination period of the drought stress seeds, so that the water potential is kept constant in the seed germination process.
5. The method of claim 2, wherein the experiment is repeated 3-5 times per 20-50 seeds in the petri dish.
6. The method of claim 1, wherein the pistacia stratiotes seeds are pretreated prior to 60 Co-gamma irradiation: selecting seeds which are not damaged and have consistent sizes, disinfecting the seeds for 20-25 min by 0.5-1% of sodium hypochlorite respectively, washing the seeds for 3-5 times by distilled water, and sucking the attached water by filter paper.
7. Use of the method according to any one of claims 1 to 6 for salt and/or drought tolerant breeding of Andrographis paniculata Nees.
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