Application of Pantoea alhagi exopolysaccharide in reduction of heavy metal cadmium enrichment in rice planting

Abstract

According to the application of the Pantoea alhagi extracellular polysaccharide in reduction of heavy metal cadmium enrichment in rice planting, oxidative stress caused by Cd stress can be relieved by applying the Pantoea alhagi extracellular polysaccharide before sowing, the Cd enrichment coefficient between soil and rice roots is reduced, toxicity of Cd to rice is relieved, and the content of Cd in rice is reduced. Meanwhile, the exopolysaccharide is further treated, so that the effect of reducing the enrichment of heavy metal cadmium in rice planting is better.

Description

Technical Field

[0001] The invention belongs to the technical field of crop cultivation, and in particular relates to a method for reducing the enrichment of heavy metal cadmium (Cd) in rice. Background Art

[0002] The main pollutants that affect the environmental quality of agricultural soil are heavy metals, among which Cd is the primary pollutant. The accumulation of Cd in plants can cause physiological damage such as chlorosis of plant leaves, metabolic disorders, reduced photosynthesis, inhibition of transpiration, and reduced enzyme activity. Heavy metal Cd can inhibit the growth and development of rice, leading to a decrease in rice yield, and eventually enter the human body through the food chain, endangering health. Rice is the main food crop in my country. It has a high absorption capacity for heavy metal Cd and can easily accumulate Cd. Rice is the main source of dietary Cd intake for the Chinese population. Therefore, reducing Cd pollution in paddy soil is of great practical significance.

[0003] Pantoeaalhagi exopolysaccharides (PAPS) are a new type of microbial biostimulant. PAPS is a secondary metabolite secreted by microorganisms into the environment. It has the characteristics of non-toxicity, biodegradability, high yield, easy separation, and short production cycle. At present, PAPS has not been used to repair heavy metal Cd pollution. Summary of the invention

[0004] Purpose of the invention: The technical problem to be solved by the present invention is to provide a method for reducing the accumulation of heavy metal cadmium in rice in view of the shortcomings of the prior art. The present invention shows that the extracellular polysaccharide of Pantoea acuminatum can be used as a biological chelator under the condition of heavy metal Cd stress, alleviate the oxidative stress caused by Cd stress, reduce the enrichment coefficient of Cd between soil and rice roots, reduce the toxicity of Cd to rice, and reduce the content of Cd in rice.

[0005] In order to solve the above technical problems, the present invention discloses an application of Pantoea aviculare exopolysaccharide in reducing the accumulation of heavy metal cadmium in rice cultivation, characterized in that the Pantoea aviculare exopolysaccharide is prepared by fermenting Pantoea aviculare, and the strain collection number of the Pantoea aviculare is CGMCC NO.15525.

[0006] Wherein, the molecular weight of the Pantoea camelthorn exopolysaccharide is not less than 1000 kDa.

[0007] Specifically, the Pantoea camellia exopolysaccharide is prepared by the following method:

[0008] (1) preparing a fermentation liquid of Pantoea camelid, adjusting the pH of the fermentation liquid to 11-12, and then adding water to dilute it;

[0009] (2) adding an excess of diatomaceous earth or perlite to the fermentation liquid obtained in step (1), mixing thoroughly, and then removing the bacterial cells by suction filtration to obtain a clarified filtrate;

[0010] (3) The filtrate obtained in step (2) is concentrated, deproteinized, ethanol precipitated, and vacuum freeze-dried to obtain extracellular polysaccharides.

[0011] Wherein, in step (1), the fermentation broth is obtained by the following steps:

[0012] 1) Pick a single colony cultured on a solid LB plate and inoculate it into a fresh LB liquid medium to obtain a seed solution;

[0013] 2) The seed liquid is transferred to the fermentation medium of Pantoea camelid polysaccharide at an inoculation rate of 8-10% v / v, the fermentation temperature is 28-32° C., the pH is controlled at 6.6-7.2, the ventilation volume is 0.8-1vvm, the stirring speed is 500-600rpm, and the fermentation is carried out for 24-28 hours to obtain the fermentation liquid of Pantoea camelid exopolysaccharide.

[0014] Preferably, the fermentation medium is composed of: sucrose 60-70 g / L, peptone 10-15 g / L, NaCl 5-7 g / L, dipotassium hydrogen phosphate 1-2 g / L, and the pH is adjusted to 6.8-7.2.

[0015] In a preferred embodiment, the seed liquid is transferred to the fermentation medium of camel thorn Pantoea polysaccharide at an inoculation rate of 10% v / v, the fermentation temperature is 30°C, the pH is controlled at 6.6-7.2, the ventilation volume is 1vvm, the stirring speed is 500rpm, and the fermentation is carried out for 24 hours to obtain the fermentation liquid of camel thorn Pantoea NX-11 extracellular polysaccharide, wherein the fermentation medium is composed of: sucrose 60g / L, peptone 10g / L, NaCl 5g / L, potassium dihydrogen phosphate 1g / L, and the pH is adjusted to 7.

[0016] Further preferably, the exopolysaccharide obtained in step (3) is further dissolved in deionized water to obtain an exopolysaccharide aqueous solution with a mass concentration of 2-3%, and then a hydrogen peroxide solution is added to the exopolysaccharide aqueous solution at a volume ratio of 2-5%, and then alcohol precipitation and freeze-drying are performed to obtain a camel thorn pantoea polysaccharide with a reduced molecular weight. Preferably, the mass concentration of the hydrogen peroxide is 30%, and alcohol precipitation and freeze-drying are performed to obtain a camel thorn pantoea polysaccharide with a reduced molecular weight.

[0017] When applied, the Pantoea acuminata exopolysaccharide is added into the rice soil once before sowing.

[0018] The amount of Pantoea aviculare exopolysaccharide added is 0.4-0.6 grams of Pantoea aviculare exopolysaccharide per kilogram of paddy field soil.

[0019] The Pantoea camellia exopolysaccharide alleviates the oxidative stress caused by cadmium stress on rice, and reduces the enrichment coefficient of Cd from soil to rice roots and the transfer coefficient of Cd from rice roots to grains.

[0020] The exopolysaccharide of Pantoea acuminata reduces the content of Cd in rice.

[0021] PAPS contains multiple negatively charged functional groups in its structure, which can chelate heavy metal ions, thereby reducing the bioavailability of heavy metals and alleviating their toxicity, and improving the growth of plants under Cd stress. Therefore, PAPS has great potential in the remediation of heavy metal Cd-contaminated soil.

[0022] Beneficial effects: The present invention proposes that the use of Pantoea thornii extracellular polysaccharides can effectively alleviate the stress of heavy metal Cd on rice growth. Under the action of Pantoea thornii extracellular polysaccharides, the chlorophyll and proline content and the activity of antioxidant enzymes in rice seedlings were significantly increased, and the malondialdehyde content was significantly reduced, thereby alleviating the oxidative stress caused by Cd stress. At the same time, the enrichment coefficient of Cd from the soil to the roots of rice was reduced, effectively limiting the enrichment of Cd in rice, thereby reducing the Cd content in rice grains. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 is the liquid chromatogram of PAPS-A and PAPS-B;

[0024] Figure 2 The changes of chlorophyll content in rice in different treatment groups 30 days after rice seedlings were transplanted;

[0025] Figure 3 The changes of malondialdehyde content in rice in different treatment groups 30 days after rice transplanting;

[0026] Figure 4 The changes of proline content in rice in different treatment groups 30 days after rice seedlings were transplanted;

[0027] Figure 5 The changes of CAT content in rice in different treatment groups 30 days after rice transplanting;

[0028] Figure 6 The changes of POD content in rice in different treatment groups 30 days after rice transplanting;

[0029] Figure 7 The changes of SOD content in rice in different treatment groups 30 days after rice transplanting;

[0030] Figure 8 is the content of heavy metal Cd in rice roots and grains in different treatment groups after rice matures;

[0031] Fig. 9 is the enrichment coefficient of Cd in different treatment groups. DETAILED DESCRIPTION

[0032] The present invention will be further described in detail below in conjunction with the accompanying drawings and specific embodiments, and the above and / or other advantages of the present invention will become more clear.

[0033] The Pantoea alhagi used in the present invention is Pantoea alhagi NX-11, with a strain deposit number of CGMCCNO.15525, and its detailed information is disclosed in CN108865951A. The rice seeds (Oryza sativassp. japonica cv. Nipponbare) used for testing were provided by Jiangsu Academy of Agricultural Sciences (i.e., Nipponbare), but the implementation of the present invention is not limited to this variety).

[0034] Example 1 Preparation of Pantoea camellia exopolysaccharide.

[0035] (1) Preparation of Fermentation Broth of Extracellular Polysaccharide from Pantoea acuminata

[0036] 1) Pick a single colony cultured on a solid LB plate and inoculate it into fresh LB liquid medium, and culture it overnight at 37°C and 200 rpm to obtain seed solution;

[0037] 2) The seed liquid was transferred to the fermentation medium of Pantoea camelidae polysaccharide at an inoculation rate of 10% v / v, the fermentation temperature was 30° C., the pH was controlled at 6.6-7.2, the ventilation volume was 1 vvm, the stirring speed was 500 rpm, and the fermentation was carried out for 24 hours to obtain the fermentation liquid of Pantoea camelidae X-11 exopolysaccharide;

[0038] The fermentation medium is composed of: 60 g / L sucrose, 10 g / L peptone, 5 g / L NaCl, and 1 g / L dipotassium hydrogen phosphate; and the pH is adjusted to 7.

[0039] (2) Preparation of extracellular polysaccharides from Pantoea acuminata

[0040] 1) adjusting the pH of the fermentation broth of Pantoea alhagi exopolysaccharide to 11-12, wherein the Pantoea alhagi used is Pantoea alhagi NX-11, and the strain collection number is CGMCC NO.15525, and then diluting it with 3 times the volume of deionized water;

[0041] 2) adding an excess amount of diatomaceous earth to the fermentation liquid obtained after the treatment in step (1), mixing thoroughly, and then filtering to remove the bacterial cells to obtain a clarified filtrate;

[0042] 3) The filtrate obtained in step 2) was concentrated to 1 / 5 of the initial volume, and the supernatant obtained after concentration was deproteinized 3 times using Sevage reagent (n-butanol: chloroform = 1:4, volume ratio); then 3 volumes of 95% ethanol were added and precipitated at 4°C overnight to obtain crude polysaccharide, and the precipitate was vacuum freeze-dried to obtain extracellular polysaccharide (1000-2000 kDa), which was recorded as PAPS-A.

[0043] The obtained extracellular polysaccharide was dissolved in deionized water to obtain a 2wt% extracellular polysaccharide aqueous solution, and then a hydrogen peroxide (30wt%) solution was added to the extracellular polysaccharide aqueous solution with a volume ratio of 2%. After alcohol precipitation and freeze-drying, the camel thorn parasite polysaccharide with reduced molecular weight was obtained, which was recorded as PAPS-B (100-1000kDa).

[0044] The average molecular weight of PAPS was determined by high performance gel permeation chromatography with multi-angle laser light scattering and refractive index analysis (HPGPC-MALLS-RI) (C.LI, D.CHEN, J.DING, et al. A Novel Hetero-Exopolysaccharide for the Adsorption of Methylene Blue from Aqueous Solutions: Isotherm, Kinetic, and Mechanism Studies [J]. J Clean Prod, 2020, 265: 121800). The chromatographic column used was: Ultrahydrogel TM 500 gel filtration column (7.8×300mm) and Ultrahydrogel TM A linear gel filtration column (7.8×300 mm) (Waters, USA) was used. The detection conditions were: 35°C, 0.6 mL / min; the mobile phase was: 0.1 M Na 2 SO 4 The weight average molecular weight of PAPS-A is 1000-2000 kDa, and the weight average molecular weight of PAPS-B is 100-1000 kDa. After further treatment, the extracellular polysaccharide PAPS-B with reduced molecular weight is obtained.

[0045] Example 2: Pantoea aconitifolia exopolysaccharide alleviates heavy metal Cd stress in rice.

[0046] (1) Cultivation of rice seedlings:

[0047] The rice seeds with full grains and uniform size were surface sterilized with 2.5% sodium hypochlorite for 10 minutes, rinsed with sterile water 10 times, and then evenly spread in a sterile culture dish with two layers of gauze, and added with an appropriate amount of sterile water. After germination at 25°C in the dark for 3 days, the seeds were placed in a culture box, and fresh nutrient solution (1 / 2MS culture medium) was replaced every three days. After two weeks of culture, rice seedlings were obtained.

[0048] (2) Soil preparation:

[0049] CK group: untreated brown soil;

[0050] Cd group: 1 mg Cd was mixed into each kg of brown soil;

[0051] Cd+PAPS1-1 (CP 1-1) group: 1 mg Cd and 0.2 g PAPS were mixed per kg brown soil;

[0052] Cd+PAPS1-2 (CP 1-2) group: 1 mg Cd and 0.4 g PAPS were mixed per kg brown soil;

[0053] Cd+PAPS1-3 (CP 1-3) group: 1 mg Cd and 0.6 g PAPS were mixed per kg brown soil;

[0054] Cd+PAPS2-1 (CP 2-1) group: 1 mg Cd and 0.2 g PAPS were mixed per kg brown soil;

[0055] Cd+PAPS2-2 (CP 2-2) group: 1 mg Cd and 0.4 g PAPS were mixed per kg brown soil;

[0056] Cd+PAPS2-3 (CP 2-3) group: 1 mg of Cd and 0.6 g of PAPS were mixed per kilogram of brown soil.

[0057] Among them, Cd+PAPS1-1, Cd+PAPS1-2 and Cd+PAPS1-3 applied PAPS-A extracted from the fermentation broth, and Cd+PAPS2-1, Cd+PAPS2-2 and Cd+PAPS2-3 applied PAPS-B.

[0058] The rice seedlings cultivated in (1) were transplanted into the soil of step (2) to obtain rice plants under different treatments, with 8 pots repeated for each treatment group, and 9 rice plants planted in each pot. Each pot was filled with 10 kg of soil, and fertilizer was supplied at a normal application level. The PAPS and Cd were applied once before rice transplanting. The rice was collected in two batches, one batch was collected 30 days after rice transplanting, and the other batch was collected after the rice matured. The rice samples collected 30 days after transplanting were used to determine the chlorophyll, proline, malondialdehyde content and antioxidant enzyme (SOD, POD, CAT) content of the plants. The samples collected after the rice matured were used to determine the Cd content in the roots and grains.

[0059] Experimental determination methods for each indicator:

[0060] Determination of chlorophyll content: 0.5 g of leaves were cut into discs and bleached with 50 mL of 80% (v / v) acetone aqueous solution at room temperature in the dark for 48 hours. Then, the suspension was centrifuged at 10,000×g for 5 min. Finally, the absorbance of the supernatant was measured at two wavelengths (645 and 663 nm), and the chlorophyll content was calculated using the following formula: C (mg / g) = 8.05×A 663 +20.29×A 645 .

[0061] Determination of malondialdehyde (MDA) content: The malondialdehyde content was determined by the thiobarbituric acid (TBA) colorimetric method. Grind 0.5 g of rice roots and 10 mL of 5% (w / v) trichloroacetic acid (TCA), homogenize and centrifuge at 12,000×g for 15 min. Then, mix the supernatant with 5 mL of 0.5% thiobarbituric acid (TBA; prepared with 20% TCA), and place the mixture in a boiling water bath for 25 minutes. After the reaction solution was cooled to room temperature, centrifuge it at 7,500×g for 5 min. Finally, the absorbance of the supernatant was measured at 450, 532 and 600 nm, and the MDA content was calculated using the following formula: C (mmol / g) = 6.45×(A 532 -A 600 )-0.56×A 450 .

[0062] Determination of proline content: The proline content was determined by the acid ninhydrin method. The rice root (0.5 g) was chopped and placed in a test tube, and 5 mL of 3% sulfosalicylic acid was added. The test tube was bathed in boiling water for 10 minutes, and then 2 mL of the supernatant was mixed with 2 mL of acetic acid and 3 mL of 2.5% ninhydrin. After mixing, the mixture was bathed in boiling water again for 40 minutes, and then extracted with 4 mL of methylbenzene. Finally, the absorbance of the extract was measured at a wavelength of 520 nm and the proline content in the rice root was calculated based on the proline standard curve.

[0063] Activity determination of antioxidant enzymes SOD, POD and CAT: 0.5 g of rice roots were homogenized in 1.5 mL of Tris-HCl buffer (pH 7.5) containing 5% sucrose and 0.1% mercaptoethanol. The homogenate was centrifuged at 10,000 × g for 20 min at 4°C. Then, the supernatant was used as a crude enzyme extract to determine the activities of antioxidant enzymes SOD, CAT and POD.

[0064] SOD enzyme activity assay: The reaction system contained 1.5 mL of 0.05 M phosphate buffer, 0.3 mL of 130 mM methionine solution, 0.3 mL of 750 μM nitro blue tetrazolium (NBT) solution, and 0.3 mL of 100 μM EDTA-Na 2 Solution, 0.3mL of 20μM lactochromium solution, 0.5mL of distilled water and 0.1mL of crude enzyme extract. The reaction was started under 4000lx light and maintained for 20min. A reaction in which phosphate buffer was used instead of crude enzyme extract for 20min under the same light intensity was used as a control. Another reaction containing phosphate buffer and incubated in the dark for 20 minutes was used as a blank. At the end of the reaction, the absorbance was measured at 560nm. One unit of SOD enzyme activity is defined as the amount of enzyme that inhibits the reduction of NBT by 50%, and the results are expressed as units per mg of protein.

[0065] POD enzyme activity assay: The reaction system contained 2.9 mL of 0.05 M phosphate buffer, 0.5 mL of 2% H 2 O 2 , 0.1 mL of 2% guaiacol and 0.1 mL of crude enzyme extract. At the end of the reaction, the absorbance was measured at 470 nm. POD activity was defined as the amount of guaiacol oxidized per minute and expressed as nanomoles per minute per mg of protein.

[0066] CAT enzyme activity assay: The reaction system contained 0.1 mL of 2% H 2 O 2 and 2 mL of 50 mM phosphate buffer (pH 7.0), and the reaction was started by adding 0.1 mL of crude enzyme extract. At the end of the reaction, the absorbance was measured at 240 nm. CAT activity was reported as H per minute. 2 O 2 The decrease was expressed as nanomoles per minute per milligram of protein.

[0067] Determination of Cd content in rice roots and grains: Wash the rice samples with deionized water three times and dry them with absorbent paper. Fresh rice samples were first sterilized in a 105°C oven for 30 minutes, and then placed in a 65°C oven to dry until the mass was constant. The dried samples were ground into powder using a grinder. Weigh 0.3 g of rice sample and put it into a digestion tube, add 10 mL of nitric acid-perchloric acid mixed solution overnight (9+1), cover and soak overnight, add a small funnel and digest on a heavy metal digestion instrument until the digestion solution is colorless and transparent or slightly yellowish, cool and wash the digestion solution into a 25 mL volumetric flask, wash the digestion tube three times with a small amount of nitric acid solution, combine the washing solution in a volumetric flask and dilute to the scale with 1% nitric acid solution, mix well; at the same time, perform a reagent blank test (GB 5009.15-20141).

[0068] The bioconcentration factor (BCF) of Cd indicates the ability of rice to accumulate Cd, that is, the ratio of the Cd content in rice to the total Cd content in the soil. The larger the value, the stronger the ability of rice to accumulate Cd. The formula is as follows: BCF = Cd grain / Cd soil Where: Cd soil and Cd grain and are the Cd concentrations in soil and rice grains (mg / kg), respectively.

[0069] The experimental results are as follows Figure 2-Figure 9 shown.

[0070] Figure 2 The results showed that compared with the CK group, the chlorophyll content in the Cd group was significantly reduced by 43.51%, indicating that Cd stress inhibits the synthesis of rice chlorophyll, thereby inhibiting the growth of rice. Compared with the Cd group, the application of different concentrations of PAPS-A and PAPS-B can significantly increase the chlorophyll content of rice (CP1-1 group increased by 28.63%, CP1-2 group increased by 47.79%, CP1-3 group increased by 57.26%, CP2-1 group increased by 49.32%, CP2-2 group increased by 56.41%, CP2-3 group increased by 58.12%). This shows that PAPS can alleviate the inhibition of chlorophyll synthesis caused by Cd stress, thereby alleviating the toxicity of Cd to rice. Among them, PAPS-B is more effective than PSPS-A at the same concentration.

[0071] The content of malondialdehyde can directly reflect the level of cell membrane peroxidation. Figure 3As shown, compared with the CK group, the malondialdehyde content in the Cd group increased significantly by 67.03%, indicating that the cell membrane of rice was damaged under Cd stress. Compared with the Cd group, the application of different concentrations of PAPS-A and PAPS-B can significantly reduce the malondialdehyde content in rice (CP1-1 group decreased by 12.27%, CP1-2 group decreased by 21.87%, CP1-3 group decreased by 25.92%, CP2-1 group decreased by 14.40%, CP2-2 group decreased by 25.39%, CP2-3 group decreased by 29.42%). This shows that PAPS can effectively alleviate the accumulation of MDA content in rice under Cd stress and reduce the membrane peroxidation of rice cells. Among them, PAPS-B is better than PSPS-A at the same concentration.

[0072] Proline is a free radical scavenger that can scavenge free radicals and maintain redox balance. Figure 4 As shown, compared with CK, the proline content in the Cd group increased significantly by 38.09, indicating that rice cells relieved the oxidative stress caused by Cd stress by increasing the proline content (CP1-1 group increased by 12.13%, CP1-2 group increased by 19.79%, CP1-3 group increased by 30.35%, CP2-1 group increased by 19.09%, CP2-2 group increased by 32.10%, and CP2-3 group increased by 36.79%). Compared with the Cd group, the application of different concentrations of PAPS-A and PAPS-B can significantly increase the proline content in rice, indicating that PAPS relieves the oxidative stress caused by Cd stress by increasing the proline content. Among them, PAPS-B has a better effect than PSPS-A at the same concentration.

[0073] Antioxidant enzymes (CAT, POD, and SOD) can convert peroxides into less toxic or harmless substances. Figure 5-7As shown in the figure, compared with the CK group, the activities of antioxidant enzymes CAT, POD and SOD in the Cd group were significantly increased (CAT increased by 37.57%, POD increased by 16.84%, SOD increased by 17.72%), indicating that rice cells alleviated the oxidative stress caused by Cd stress by increasing the activity of antioxidant enzymes. Compared with the Cd group, the application of PAPS with different concentrations and molecular weights can significantly increase the activities of antioxidant enzymes CAT, POD and SOD in rice, indicating that PAPS can further increase the antioxidant enzymes CAT (CP1-1 group increased by 17.96%, CP1-2 group increased by 39.42%, CP1-3 group increased by 48.72%, CP2-1 group increased by 30.65%, CP2-2 group increased by 47.60%, CP2-3 group increased by 55.14%), POD (CP1-1 group increased by 9.02%, CP1-2 group increased by 39.42%, CP1-3 group increased by 48.72%, CP2-1 group increased by 30.65%, CP2-2 group increased by 47.60%, CP2-3 group increased by 55.14%), and POD (CP1-1 group increased by 9.02%, CP1-3 group increased by 48.72%, CP2-1 group increased by 30.65%, CP2-2 group increased by 47.60%, CP2-3 group increased by 55.14%). -2 group increased by 16.60%, CP1-3 group increased by 21.11%, CP2-1 group increased by 11.88%, CP2-2 group increased by 20.16%, CP2-3 group increased by 24.37%) and SOD (CP1-1 group increased by 18.52%, CP1-2 group increased by 30.06%, CP1-3 group increased by 39.29%, CP2-1 group increased by 26.55%, CP2-2 group increased by 37.13%, CP2-3 group increased by 44.04%) activities to alleviate the oxidative stress caused by Cd stress. Among them, PAPS-B is more effective than PSPS-A at the same concentration.

[0074] Example 3 Effects of PAPS of different concentrations and molecular weights on Cd content and Cd enrichment coefficient in rice roots and grains under Cd stress.

[0075] like Figure 8 As shown in the results, the application of PAPS with different concentrations and molecular weights could significantly reduce the Cd content in rice roots and grains.

[0076] The Cd contents in rice grains of different treatment groups are as follows:

[0077] CK group: 0.5095mg / kg,

[0078] The Cd group was 3.2723 mg / kg.

[0079] The Cd+PAPS1-1 group was 2.5595 mg / kg.

[0080] The Cd+PAPS1-2 group was 2.3485 mg / kg.

[0081] The Cd+PAPS1-3 group was 2.2148 mg / kg,

[0082] The Cd+PAPS2-1 group was 2.4467 mg / kg.

[0083] The Cd+PAPS2-2 group was 2.1128 mg / kg.

[0084] The Cd+PAPS2-3 group was 2.0128 mg / kg.

[0085] The Cd contents in rice grains of different treatment groups are as follows:

[0086] CK group: 0.039mg / kg,

[0087] The Cd group was 0.4275 mg / kg.

[0088] The Cd+PAPS1-1 group was 0.2573 mg / kg.

[0089] The Cd+PAPS1-2 group was 0.2213 mg / kg.

[0090] The Cd+PAPS1-3 group was 0.2215 mg / kg.

[0091] The Cd+PAPS2-1 group was 0.2285 mg / kg.

[0092] The Cd+PAPS2-2 group was 0.198 mg / kg.

[0093] The Cd+PAPS2-3 group was 0.1912 mg / kg.

[0094] like Fig. 9 As shown in the figure, applying different concentrations of PAPS-A and PAPS-B can significantly reduce the enrichment coefficient of Cd. This shows that PAPS can effectively inhibit the enrichment of Cd in rice soil and reduce the Cd content in rice grains. Among them, PAPS-B is more effective than PSPS-A at the same concentration.

[0095] Taking all factors into consideration, applying 0.4 g / kg PAPS-B can significantly reduce the accumulation of heavy metal Cd in brown soil rice.

[0096] The present invention provides the application of Pantoea camelidae exopolysaccharide in reducing the accumulation of heavy metal cadmium in rice cultivation. There are many methods and ways to implement the technical solution. The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention. All components not specified in this embodiment can be implemented by existing technologies.

Claims

1. Application of Pantoea acuminatum exopolysaccharide in reducing the accumulation of heavy metal cadmium in rice cultivation, characterized in that: The extracellular polysaccharide of Pantoea acuminata is prepared by fermenting Pantoea acuminata, and the strain collection number of Pantoea acuminata is CGMCCNO.15525.

2. The use according to claim 1, characterized in that: The molecular weight of the Pantoea camelthorn exopolysaccharide is not less than 1000 kDa.

3. The use according to claim 1, characterized in that: The exopolysaccharide of Pantoea acuminata is prepared by the following method: (1) preparing a fermentation broth of Pantoea camelid, adjusting the pH of the fermentation broth to 11-12, and then adding water to dilute it; (2) adding diatomaceous earth or perlite to the fermentation liquid obtained in step (1), mixing thoroughly, and then removing the bacterial cells by suction filtration to obtain a clarified filtrate; (3) The filtrate obtained in step (2) is concentrated, deproteinized, ethanol precipitated, and vacuum freeze-dried to obtain extracellular polysaccharides.

4. The use according to claim 3, characterized in that: In step (1), the fermentation broth is obtained by the following steps: 1) Pick a single colony cultured on a solid LB plate and inoculate it into fresh LB liquid medium to obtain seed solution; 2) The seed liquid is transferred to the fermentation medium of camel thorn Pantoea polysaccharide at an inoculation rate of 8-10% v / v, the fermentation temperature is 28-32°C, the pH is controlled at 6.6-7.2, the ventilation volume is 0.8-1 vvm, the stirring speed is 500-600 rpm, and the fermentation is carried out for 24-28 hours to obtain the fermentation liquid of camel thorn Pantoea exopolysaccharide.

5. The use according to claim 4, characterized in that: The fermentation medium is composed of: 60-70 g / L sucrose, 10-15 g / L peptone, 5-7 g / L NaCl, 1-2 g / L potassium hydrogen phosphate, and the pH is adjusted to 6.8-7.

2.

6. The use according to claim 3, characterized in that: The extracellular polysaccharide obtained in step (3) is further dissolved in deionized water to obtain an extracellular polysaccharide aqueous solution with a mass concentration of 2-3%, and then a hydrogen peroxide solution is added to the extracellular polysaccharide aqueous solution at a volume ratio of 2-5%, and then alcohol precipitation and freeze-drying are performed to obtain a camel thorn pantothecoides polysaccharide with a reduced molecular weight.

7. The use according to claim 1, characterized in that: The Pantoea acuminata exopolysaccharide is added into the rice soil once before sowing.

8. The use according to claim 7, characterized in that: The amount of Pantoea acuminata extracellular polysaccharide added is 0.4-0.6 grams of Pantoea acuminata extracellular polysaccharide per kilogram of paddy field soil.

9. The use according to claim 1, characterized in that: The extracellular polysaccharide of Pantoea acuminatum alleviates the oxidative stress caused by cadmium stress on rice and reduces the enrichment coefficient of Cd between soil and rice roots.

10. The use according to claim 8, characterized in that: The exopolysaccharide of Pantoea acuminata reduces the content of Cd in rice.

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