CN113897320A - Halophyte with salt-tolerant growth-promoting function and application thereof - Google Patents
Halophyte with salt-tolerant growth-promoting function and application thereof Download PDFInfo
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Abstract
The invention discloses a Zeshu salina strain JP11 with salt-resistant and growth-promoting functions, which is preserved in China general microbiological culture Collection center (CGMCC) on 9, 14 and 2021 with the preservation number of CGMCC No. 23407. The invention also discloses application of the Zyrhengzhi salt pan bacteria in relieving inhibition of salt stress on rice growth and a liquid culture medium suitable for fermentation of the Zyrhengzhi salt pan bacteria. Experiments prove that the Zhang Yantian fungus JP11 has multiple growth promoting characteristics such as potassium dissolving, phosphorus dissolving and phosphorus dissolving, high salt tolerance and wide salt tolerance spectrum, the fungus solution prepared by JP11 can improve the salt tolerance growth promoting capability of rice under salt stress, accelerate the growth of seedlings and increase the biomass, thereby promoting the germination of rice and the growth of seedlings, meanwhile, the JP11 provided by the invention can increase the content of soluble potassium in soil by 25.4% and the content of soluble phosphorus in soil by 45.9%, effectively improve the utilization rate of soil and provide excellent strain resources for the restoration of saline-alkali farmland soil.
Description
Technical Field
The invention relates to a salt pan bacterium and application thereof, in particular to a Zeshu salt pan bacterium JP11 with salt-tolerant growth-promoting function and application thereof in relieving inhibition of salt stress on rice growth, and belongs to the technical field of agricultural microorganisms.
Background
Soil salinization has become a serious threat to the agroecological system. 15 to 50 percent of irrigated land in the world suffers salinization, and the inhibition effect of the salinization on the plant growth severely restricts the production of crops. The saline-alkali land treatment is a major difficult problem to overcome urgently all over the world. Besides the traditional physical and chemical methods, biological treatment has become a new direction for saline-alkali soil improvement. Wherein, the saline-alkali soil is treated by functional microorganisms, so that the comprehensive benefits of environment, ecology and economy can be integrated, the saline-alkali soil is effectively improved, and the crop yield is increased. Therefore, the method has important significance for collecting and perfecting the efficient and practical soil functional microorganism resources.
The saline-alkali soil special habitat with high salt and poor nutrition contains a plurality of rich microbial resources which are suitable for the special habitat, in particular plant growth promoting strains with high salt tolerance. By excavating the native salt-tolerant growth-promoting functional microorganisms of the saline-alkali soil and utilizing the growth-promoting functions of phosphate dissolving, potassium dissolving, phosphorus dissolving, nitrogen fixation and the like, the environment of plant flora can be effectively improved, the quick-acting nutrient elements in the soil are increased, the improvement and restoration of the saline-alkali soil are accelerated, and the growth of crops is promoted, so that the utilization rate of the saline-alkali soil is improved.
The salt-tolerant growth-promoting bacteria provide nutrient substances for plants through interaction with the plants, and can effectively relieve the toxic action of salt ions in the environment on the plants. However, the prior art has less research on salt-tolerant growth-promoting strains, and the strain resources applicable to actual production are still poor. The high-salt-tolerance culturable microorganisms with multiple growth promoting functions of potassium dissolving, phosphorus dissolving and the like can particularly survive in a high-salinity environment, can effectively improve the utilization rate of saline-alkali soil and promote the growth of crops, and the application of the Zuishu saline-alkali soil bacteria in relieving the inhibition of salt stress on the growth of the crops is not reported.
Disclosure of Invention
Aiming at the defects of the prior art, the invention aims to provide a Zeshu salina strain with salt-tolerant growth-promoting function and application thereof in relieving the inhibition of salt stress on rice growth.
The invention relates to a Zeshu salina strain with salt-resistant growth-promoting function, which is characterized in that: the strain is named as Zephysalis zeshuii JP11, and the strain is preserved in China general microbiological culture Collection center (address: Beijing Shangyang district Beichen Xilu No.1 Hospital No. 3) on 9-14 th of 2021 with the preservation number of CGMCC No. 23407.
The Zhang Shu Yan Tian fungus with salt-resistant growth-promoting function has the biological characteristics that: the shape of the bacterial cells is short rod-shaped, single, the cell size is (0.5-0.8 μm) × (1.0-1.3 μm), and no spore is generated. When the solid culture is carried out at the temperature of 28 ℃, the bacterial colony is regular and round, small, milky white, opaque, smooth and moist in surface, easy to pick and neat in edge; the physiological and biochemical characteristics are as follows: the growth temperature is 28-34 ℃, the pH value is 6.0-8.0, the salt concentration is 6-14%, the salt tolerance range is 1-18%, and the characteristics of potassium dissolution, phosphorus dissolution and phosphorus dissolution are achieved.
The result of the 16S rRNA gene sequence determination of the Hizikia salina strain JP11 shows that the gene length is 1385bp, and the corresponding nucleotide sequence is shown as SEQ ID NO. 1.
Strain JP11 of the present invention was determined to be a strain of saxifraga zeshunii (Salinicola zeshunii) by alignment and phylogenetic analysis using the american Center for Biotechnology Information (NCBI) BLASTN program.
Selecting 16S rDNA sequences of 20 sequences with high homology as reference objects, and constructing a phylogenetic tree between the strain JP11 and the reference strain by using a Mega 7 software through a proximity method (neighbor-Joining). In the clade, strain JP11 forms a separate intra-cluster clade with the zearales salina model strain salicorna zeshunii N4 (fig. 7). The physiological and biochemical results were generally consistent with those of the model strain Salinicola zeshunii N4, as detailed in Table 1.
TABLE 1 comparison of the physio-biochemical characteristics of Strain JP11 with Salinicola zeshunii N4
Note: "+" indicates positive strain; "-" indicates that the strain is negative.
The liquid culture medium for observing the shape of the thallus comprises the following components: each 1000mL of distilled water contained 10g of peptone, 5g of yeast powder and 80g of sodium chloride, and the pH was adjusted to 8.0.
The solid culture medium for observing the shape of the thallus comprises the following components: each 1000mL of distilled water contains 10g of peptone, 5g of yeast powder, 80g of sodium chloride and 15g of agar powder, and the pH is adjusted to 8.0.
The invention optimizes a liquid culture medium suitable for the fermentation of the Zyguiziqi salina strain JP11, and is characterized in that: the fermentation medium comprises the following components: every 1000mL of tap water contains 40g of glucose, 10g of yeast powder, 20g of peptone, 80g of sodium chloride, 2g of potassium sulfate, 0.01g of calcium chloride, 1g of potassium feldspar and 2g of lecithin, and the pH is adjusted to 7.5-8.0.
The basic method for breeding the Zeshu salina strain JP11 with the salt-tolerant growth-promoting function comprises the following steps:
collecting suaeda salsa root soil from saline-alkali soil in a natural protection area of yellow river delta, oscillating and uniformly mixing to prepare a soil suspension, performing gradient dilution and coating on an LB (Luria Bertani) flat plate containing 8% NaCl, performing static culture at the temperature of 28-30 ℃, selecting a single colony on the LB culture flat plate with the same salt concentration, performing purification twice to obtain a strain, and performing frozen storage and preservation on glycerol.
The invention discloses application of Zeshu salina bacteria with salt-tolerant growth-promoting functions in relieving inhibition of salt stress on rice growth.
Wherein: the salt stress refers to the condition of farmland soil with the salt concentration mass ratio not less than 3 per thousand in the soil.
The method related to the application is as follows:
(1) inoculating the activated strain JP11 seed solution into the fermentation liquid culture medium of the Zygus lucidus strain according to the inoculation amount of 2-5% of the volume ratio, and culturing for 48-60h at the temperature of 28-30 ℃ and under the condition of 180rpm to obtain a bacterial suspension containing the Zygus lucidus strain;
(2) soaking rice seeds in tap water for 24 +/-2 hours, taking full seeds with consistent sizes, and uniformly arranging the seeds in a flowerpot filled with equivalent sterilized nutrient soil poured with 7 per mill of NaCl solution; diluting the fermented ZE-SHI salina fungus suspension with different gradients to obtain 1-5 × 107cfu/mL、1～5×108cfu/mL and 1-5X 109cfu/mL three concentrations of the Zymus salina JP11 bacterial suspension; respectively irrigating three kinds of bacteria liquid with the same amount and different concentrations into the flowerpot, and taking the inactivated bacteria liquid as a reference; and pouring 7 per mill NaCl solution into all the flowerpots; irrigating with 7% NaCl solution every other day; and observing the growth condition of the rice in the seedling stage after ten days, and detecting the content of soluble potassium and soluble phosphorus in the soil around the roots of the rice.
The results show that: the growth vigor of the rice seedlings applying the JP11 bacterial liquid is obviously superior to that of a control group, and the bacterial strain JP11 is proved to be capable of obviously relieving the inhibition effect of salt stress on the growth of the rice, meanwhile, the content of soluble potassium in soil is increased by 25.4%, and the content of soluble phosphorus in the soil is increased by 45.9%.
Furthermore, the invention discloses an application of the Zygroshima halophyte with the salt-tolerant growth-promoting function in increasing the content of soluble potassium in soil, wherein the content of the soluble potassium in the soil is increased by 25.4%. The invention relates to an application of Zeshu salina bacteria with salt-tolerant growth-promoting function in increasing the content of soluble phosphorus in soil, wherein the content of soluble phosphorus in soil is increased to 45.9%, and the phosphorus dissolution index are respectively 2.4 and 2.7.
The invention discloses a Zyrhizi halophyte JP11 with salt-tolerant growth-promoting function, which has the characteristics of high salt tolerance, potassium dissolution, phosphorus dissolution and the like, has a wide salt-tolerant range, can keep good growth under the condition of 1-18% salt concentration, and has great potential for being applied to saline-alkali soil remediation and agricultural production. The innovation of the invention is that a salt-tolerant strain with multiple growth promoting functions is obtained through multiple rounds of screening, and under high salt stress, the strain can relieve the inhibition effect of the salt stress on the growth of rice, promote the growth of the rice, and simultaneously obviously increase the content of soluble potassium and soluble phosphorus in soil by 25.4% and 45.9% respectively. The invention provides the Zedhair salt pan strain JP11 and also provides a liquid culture medium suitable for the fermentation of the Zedhair salt pan strain JP11, a large amount of Zedhair salt pan strain JP11 with salt-tolerant growth-promoting function can be rapidly obtained by using the culture medium, the salt-tolerant growth-promoting strain provides excellent strain resources for the soil remediation of saline-alkali farmland, and the strain and the bacterial liquid thereof have higher agricultural application prospect and economic value.
Drawings
The Zygroshima halophyte (Salinicola zeshuni) JP11 of the invention has been deposited in China general microbiological culture Collection center (CGMCC) at 9-14.2021 with the deposit number of CGMCC No. 23407.
FIG. 1 shows the microscopic cell morphology of the Hizikia salina strain JP 11.
FIG. 2 is the colony morphology of Hizikia salina JP11 on LB solid medium.
FIG. 3 is a graph showing the growth curve of Zymus salina strain JP11 under different salt concentration conditions.
FIG. 4 is a graph showing the effect of potassium-releasing of Zuishu salina strain JP 11.
FIG. 5 is a graph showing the effect of phosphate solubilizing (organic phosphorus) by Zhang Kshisha salina strain JP 11.
FIG. 6 is a graph showing the effect of solubilizing phosphorus (inorganic phosphorus) by Zuishu salina strain JP 11.
FIG. 7 is a phylogenetic tree between Zephyr salina JP11 and a reference strain.
FIG. 8 is a schematic diagram of the growth promoting effect of Zuishu salina strain JP11 on rice seedling stage.
FIG. 9 is a statistical chart of the growth heights of rice seedlings.
Detailed Description
The present invention will be described in detail with reference to the following detailed drawings and examples. The following examples are only preferred embodiments of the present invention, and it should be noted that the following descriptions are only for explaining the present invention and not for limiting the present invention in any form, and any simple modifications, equivalent changes and modifications made to the embodiments according to the technical spirit of the present invention are within the scope of the technical solution of the present invention.
In the following examples, materials, reagents and the like used were obtained commercially unless otherwise specified.
Example 1 screening of salt-resistant book-dwelling saltpan bacteria
(1) Collecting root soil samples of saline-alkali tolerant plants such as suaeda salsa, reed and the like from saline-alkali soil in natural protection area of yellow river delta, filling the root soil samples into a clean sampling bag, making a mark, placing the sample in an ice box, bringing the sample back to a laboratory, and storing the sample at-20 ℃ for later use. Weighing 1g of soil sample in a sterile super clean bench, placing the soil sample in a 100mL triangular flask containing glass beads and 20mL sterile water, and culturing for 1 hour in a shaking incubator at 28 ℃ and 180rpm to fully break up and mix the soil sample uniformly. Respectively sucking 0.5mL of soil suspension into 4.5mL of sterile water under aseptic condition, mixing, and sequentially diluting in gradient to obtain 10-1、10-2、10-3Respectively sucking 0.2mL of sample solutions with different dilutions, and coating the sample solutions on salt-tolerant screening solidAnd (3) inversely culturing the cells on a culture medium plate in a constant-temperature incubator at 28 ℃ for 2-4 days until obvious single colonies are generated. And respectively picking single colonies, transferring the single colonies to the same solid medium flat plate, sequentially numbering, culturing, and correspondingly obtaining pure culture strain single colonies after twice transferring and scribing.
(2) And (3) picking a single colony by using an inoculating loop, transferring the single colony into a test tube filled with 5mL of salt-resistant screening liquid culture medium, carrying out shake culture for 24h under the conditions of 28 ℃ and 180rpm, and respectively carrying out strain preservation and physiological and biochemical analysis. The strain preservation method adopts a glycerol tube freezing preservation method, 200 mu L of glycerol and 800 mu L of bacterial liquid are respectively added into a freezing preservation tube, and the mixture is uniformly mixed and then placed in an ultra-low temperature refrigerator at minus 80 ℃ for preservation.
Among the screened strains, one strain is in the form of short rod, has a cell size of (0.5-0.8 μm) × (1.0-1.3 μm), and does not produce spores. When the solid culture is carried out at the temperature of 28 ℃, the bacterial colony is regular and round, small, milky white, opaque, smooth and moist in surface, easy to pick and neat in edge; the growth temperature is 28-34 ℃, the pH value is 6.0-8.0, the salt concentration is 6-14%, the salt tolerance range is 1-18%, and the characteristics of potassium dissolution, phosphorus dissolution and phosphorus dissolution are achieved. This strain is numbered as JP11 and was initially identified as the selected strain of the invention.
The composition of the salt-tolerant screening solid culture medium is as follows: every 1000mL of distilled water contains 10g of peptone, 5g of yeast powder, 80g of sodium chloride and 15g of agar powder, and the pH is adjusted to 7.5-8.0.
The composition of the salt-tolerant screening liquid culture medium is as follows: each 1000mL of distilled water contains 10g of peptone, 5g of yeast powder and 80g of sodium chloride, and the pH is adjusted to 7.5-8.0.
Example 2 morphological Observation and physiological and biochemical characterization of Strain JP11
The morphology of strain JP11 was observed using an oil mirror of a nikon inverted microscope.
The culture temperature of the physiological and biochemical characteristic identification test of the strain JP11 is set to be 28 ℃. Meanwhile, the strain JP11 is analyzed for the optimum temperature, the optimum growth pH value and the optimum salt concentration when growing on an LB solid culture medium.
The biological characteristics of strain JP11 are: rod-shaped, single, cell-sized (0.5 μm-0.8 μm) × (1.0 μm-1.3 μm), no spores were produced (FIG. 1). The bacterial colony is regularly round, protruding, milky white, opaque, smooth and moist in surface, easy to pick and neat in edge when cultured at the temperature of 28 ℃ (figure 2).
The physiological and biochemical characteristics of the strain JP11 are as follows: gram stain is negative, aerobic, positive in catalase, urease, phenylalanine deaminase, methyl red test and urea hydrolysis test, and negative in starch hydrolysis, gelatin hydrolysis and chitin hydrolysis test. The optimum growth temperature is 28-34 ℃, the optimum growth pH value is 6.0-8.0, the optimum salt concentration is 6-14%, and the salt tolerance range is 1-18% (figure 3).
The result of measuring the 16S rRNA gene sequence of the strain disclosed by the invention by JP11 shows that the gene length is 1385bp, and the corresponding nucleotide sequence is shown as SEQ ID NO. 1.
Strain JP11 of the present invention was determined to be a strain of saxifraga zeshunii (Salinicola zeshunii) by alignment and phylogenetic analysis using the american Center for Biotechnology Information (NCBI) BLASTN program.
Selecting 16S rDNA sequences of 20 sequences with high homology as reference objects, and constructing a phylogenetic tree between the strain JP11 and the reference strain by using a Mega 7 software through a proximity method (neighbor-Joining). In the clade, strain JP11 forms a separate intra-cluster clade with the model strain salicorna zeshunii N4 of salicornia zeshunii (fig. 7). The physiological and biochemical results were generally consistent with those of the model strain Salinicola zeshunii N4, as detailed in Table 1.
TABLE 1 comparison of the physio-biochemical characteristics of Strain JP11 with Salinicola zeshunii N4
Note: "+" indicates positive strain; "-" indicates that the strain is negative.
The liquid culture medium for observing the shape of the thallus comprises the following components: each 1000mL of distilled water contained 10g of peptone, 5g of yeast powder and 80g of sodium chloride, and the pH was adjusted to 8.0.
The solid culture medium for observing the shape of the thallus comprises the following components: each 1000mL of distilled water contains 10g of peptone, 5g of yeast powder, 80g of sodium chloride and 15g of agar powder, and the pH is adjusted to 8.0.
Example 3 analysis of growth promoting effects of the Strain JP11 on Potassium solubilization, phosphorus solubilization and phosphorus solubilization
Transferring the single bacterial colony of the strain JP11 obtained by separation into a test tube filled with 5mL of liquid culture medium, carrying out shake culture for 24h under the conditions of 28 ℃ and 180rpm, taking 5 mu L of seed liquid to be inoculated on a silicate bacteria culture medium plate, carrying out culture for 3 days in a constant temperature incubator at 28 ℃, and observing whether a transparent oil drop-shaped bacterial colony appears on the silicate bacteria culture medium plate. The result shows that after the strain JP11 is cultured in a silicate bacteria solid medium for 2 days, transparent oil drop-shaped colonies grow on the plate, and the strain JP11 is capable of dissolving potassium feldspar and has the potassium dissolving property (figure 4).
Transferring the single bacterial colony of the strain JP11 obtained by separation into a test tube filled with 5mL of liquid culture medium, carrying out shake culture for 24h under the conditions of 28 ℃ and 180rpm, taking 5 mu L of seed liquid to be spotted on a Monkina organophosphorus solid culture plate, carrying out culture in a constant temperature incubator at 28 ℃, and observing whether a degradation transparent ring exists around the bacterial colony. The result shows that after the bacterial strain JP11 is cultured in a Monkina organic phosphorus solid culture medium for 2 days, a clear degradation ring appears around a bacterial colony, the bacterial colony is continuously cultured, the degradation ring is obviously increased, and the bacterial strain JP11 can rapidly dissolve organic phosphorus and has strong phosphate solubilizing property (Table 2 and figure 5).
The single colony of the strain JP11 obtained by separation is transferred to a test tube filled with 5mL of liquid culture medium, and is subjected to shaking culture for 24h under the conditions of 28 ℃ and 180rpm, 5 mu L of seed liquid is taken and is spotted on a Monkina inorganic phosphorus solid culture plate, and the culture is carried out in a constant temperature incubator at 28 ℃. After 6 days of culture, clear degradation circles were generated around the colonies, indicating that the strain JP11 can dissolve inorganic phosphorus and has phosphorus-dissolving properties (FIG. 6 and Table 3).
TABLE 2 analysis of the phosphate solubilizing Properties of the Strain JP11
| Bacterial strains | Diameter (D)/mm of phosphate solubilizing ring | Diameter (d)/mm of bacterial colony | Phosphorus index (D/D) |
| JP11 | 24 | 10 | 2.4 |
TABLE 3 phosphorus solubilization analysis of Strain JP11
| Bacterial strains | Diameter (D)/mm of phosphorus dissolving ring | Diameter (d)/mm of bacterial colony | Index of dissolved phosphorus (D/D) |
| JP11 | 8 | 3 | 2.7 |
The components and the concentrations of the silicate bacteria culture medium are as follows: 5g L-1Sucrose, 0.5g L-1(NH4)2SO4、0.5g L-1Yeast extract, 0.3g L-1MgSO4.7H2O、2g L-1Na2HPO4、0.03g L-1FeSO4.7H2O、0.03g L- 1MnSO4.H2O、2g L-1Potash feldspar, 15g L-1Agar.
The Monkina organophosphorus culture medium comprises the following components in concentration: 10g L-1Glucose, 0.5g L-1(NH4)2SO4、0.3g L-1NaCl、0.3g L-1MgSO4.7H2O、0.03g L-1MnSO4.H2O、0.3g L-1KCl、0.03g L- 1FeSO4.7H2O、2.0g L-1Lecithin, 15g L-1Agar.
The Monkina inorganic phosphorus culture medium comprises the following components in percentage by concentration: 10g L-1Glucose, 0.5g L-1(NH4)2SO4、0.3g L-1NaCl、0.3g L-1MgSO4.7H2O、0.03g L-1MnSO4.H2O、0.3g L-1KCl、0.03g L- 1FeSO4.7H2O、5.0g L-1Ca3(PO4)2、15g L-1Agar.
Example 4 identification method of 16S rRNA Gene of Strain JP11
The total genomic DNA of the isolated and purified strain JP11 was extracted according to the instructions of the BioTeKe bacterial genome extraction kit. The extracted total genomic DNA was detected by electrophoresis on a 1% agarose gel at 175V for 20 minutes. The 16S rRNA gene of the isolated strain was amplified using the forward primer 27F (5 '-3': AGAGAGTTTGATCCTGGCTCAG) and the reverse primer 1492R (5 '-3': GGTTACCTTGTTACGACTT), and the PCR reaction system and conditions are shown in Table 4 and Table 5:
TABLE 4 16S rRNA Gene PCR reaction System of purified Strain JP11
| Reagent | Volume of |
| 10×Ex Taq Buffer | 5μL |
| dNTP(10mM) | 1μL |
| 27F primer(25μM) | 1μL |
| 1492R primer(25μM) | 1μL |
| ExTaq DNA Polymerase | 0.5μL |
| Template DNA | 0.5μL |
| dH2O | 41μL |
TABLE 5PCR reaction Programming set-ups
| Step (ii) of | Temperature of | Time | Number of cycles |
| Denature | 95℃ | 5min | 1 |
| Denature | 95℃ | 30s | 30 |
| Anneal | 55℃ | 30s | 30 |
| |
72℃ | 90s | 30 |
| |
72℃ | 10min | 1 |
| Halt reaction | 4℃ |
The PCR product was detected by electrophoresis on a 1% agarose gel at 175V for 20 minutes. Using a gel imager, the PCR of the 16S rRNA gene was observed near the position of the 1.5kbp Marker band, and after the desired band was excised, the PCR product was purified and recovered according to the instructions of the agarose gel recovery kit. The purified and recovered PCR product was sent to a sequencing company for sequencing.
The result of measuring the 16S rRNA gene sequence of the strain JP11 shows that the gene length is 1385bp, and the corresponding nucleotide sequence is shown as SEQ ID NO. 1.
The 16S rRNA gene sequences obtained by sequencing were submitted to NCBI database (http:// www.ncbi.nlm.nih.gov) for BLASTN alignment, and the results showed that the 16S rRNA of this strain had 99% similarity to the K.lucidum Salinicola zeshuniii N4. Selecting 16S rDNA sequences of 20 sequences with high homology as reference objects, and constructing a phylogenetic tree between the strain JP11 and the reference strain by using a Mega 7 software through a proximity method (neighbor-Joining). In the evolutionary tree, strain JP11 formed a separate intra-cluster evolutionary branch with the zea salina model strain sali zeani N4 (fig. 7), and it was determined that strain JP11 according to the present invention was a strain of zea salina (Salinicola zeani) and was therefore named zea salina JP 11.
Example 5 application method of strain JP11 in relieving inhibition effect of salt stress on rice growth
(1) Selecting strains: zezu salina strain JP 11.
(2) Activating strains: inoculating the strain into a solid plate culture medium, and performing static culture at 28-30 ℃ for 24-48 h for later use.
(3) Seed culture: and (3) selecting the bacterial colony in the step (2), inoculating the bacterial colony into a liquid seed culture medium containing 5mL, and culturing for 24-30 h at the temperature of 28-30 ℃ and under the condition of 180 rpm.
(4) Fermentation culture: inoculating the seed solution into a triangular flask (500mL) containing 100mL of fermentation medium at an inoculation amount of 2%, and culturing at 28-30 ℃ and 180rpm for 48h to obtain a bacterial suspension containing the Zymus lucidus salina.
(5) Relieving salt stress on riceGrowth inhibition and detection of soluble potassium and phosphorus in soil: soaking rice seeds in tap water for 24 +/-2 h, taking full seeds with consistent sizes, and uniformly arranging the seeds in flowerpots (30cm multiplied by 24cm multiplied by 9cm) filled with equal amount of sterilized nutrient soil irrigated with 7 per mill NaCl solution; diluting the fermented ZE-SHI salina fungus suspension with different gradients to obtain 1-5 × 107cfu/mL、1～5×108cfu/mL and 1-5X 109cfu/mL three concentrations of the Zymus salina JP11 bacterial suspension; respectively irrigating three kinds of bacteria liquid with the same amount and different concentrations into the flowerpot, and taking the inactivated bacteria liquid as a reference; and pouring 7 per mill NaCl solution into all the flowerpots; irrigating with 7% NaCl solution every other day; and observing the growth condition of the rice in the seedling stage after ten days, and detecting the content of soluble potassium and soluble phosphorus in the soil around the roots of the rice. Three replicates of each treatment were placed with 40 rice seeds per replicate.
The results show that the seedling growth of the experimental group is obviously better than that of the control group under the salt stress condition of continuously irrigating the rice with 7 per mill NaCl solution, and the bacterial strain JP11 is proved to be capable of obviously relieving the inhibition effect of the salt stress on the growth of the rice, wherein the concentration of the applied bacterial solution is 1-5 multiplied by 108The growth vigor of cfu/mL rice seedlings was optimal (FIG. 8). When the seedlings grow for 15 days, only 20 percent of the seedlings in the control group are higher than 20cm, and the seedlings in the experimental group are (1-5 multiplied by 10)8cfu/mL) was greater than 20cm in height (FIG. 9). The physical and chemical indexes of the soil around the rice roots are determined (table 6), the content of soluble potassium and soluble phosphorus in the soil around the rice roots added with the JP11 bacterial solution is obviously increased compared with that of a control group, the content of soluble potassium in the soil is increased by 25.4 percent as much as possible, and the content of soluble phosphorus in the soil is increased by 45.9 percent as much as possible.
TABLE 6 detection results of soluble phosphorus and soluble potassium in soil around rice roots
The strain activation solid medium comprises the following components: every 1000mL of distilled water contains 10g of peptone, 5g of yeast powder, 80g of sodium chloride and 15g of agar powder, and the pH value is adjusted to 8.0;
the liquid seed culture medium comprises the following components: every 1000mL of distilled water contains 10g of peptone, 5g of yeast powder and 80g of sodium chloride, and the pH is adjusted to 8.0;
the optimal fermentation medium composition of the above mentioned zephyr salina strain JP11 is: every 1000mL of tap water contains 40g of glucose, 10g of yeast powder, 20g of peptone, 80g of sodium chloride, 2g of potassium sulfate, 0.01g of calcium chloride, 1g of potassium feldspar and 2g of lecithin, and the pH is adjusted to 7.5-8.0.
Sequence listing
<110> Shandong province academy of agricultural sciences
<120> halophyte having salt-tolerant growth-promoting function and application thereof
<141> 2021-11-11
<160> 1
<210> 1
<211> 1385
<212> DNA
<213> Halfway bacterium (Salinicola sp.)
<221> nucleotide sequence of 16S rRNA gene of Hizikia salina JP11
<222>（1）…（1385）
<400> 1
cgaaggttaa gctaaccact tctggtgcag tccactccca tggtgtgacg ggcggtgtgt 60
acaaggcccg ggaacgtatt caccgtgaca ttctgattca cgattactag cgattccgac 120
ttcacggagt cgagttgcag actccgatcc ggactgaggc cggctttctg ggattcgctc 180
cacctcgcgg tctcgcaacc ctttgtaccg gccattgtag cacgtgtgta gccctacccg 240
taagggccat gatgacttga cgtcgtcccc accttcctcc ggtttgtcac cggcagtctc 300
cttagagttc ccaccattac gtgctggcaa ataaggacaa gggttgcgct cgttacggga 360
cttaacccaa catttcacaa cacgagctga cgacagccat gcagcacctg tcactgcgct 420
cccgaaggca ctaaggtatc tctaccaaat tcgcaggatg tcaagggtag gtaaggttct 480
tcgcgttgca tcgaattaaa ccacatgctc caccgcttgt gcgggccccc gtcaattcat 540
ttgagtttta accttgcggc cgtactcccc aggcggtcga cttatcgcgt taactgcgcc 600
actaagtcct taaaggtccc aacggctagt cgacatcgtt tacggcgtgg actaccaggg 660
tatctaatcc tgtttgctac ccacgctttc gcacctcagt gtcagtgtca ggccagaagg 720
ccgccttcgc cactggtatt cctcccgatc tctacgcatt tcaccgctac accgggaatt 780
ctaccttcct ctcctgcact ctagcctggc cgttccggat gccgttccca ggttgagccc 840
ggggctttca caaccggcgt gccaagccac ctacgcgcgc tttacgccca gtaattccga 900
ttaacgctcg caccctccgt attaccgcgg ctgctggcac ggagttagcc ggtgcttctt 960
ctgcgagtga tgtccttcct gatgggtatt aaccaccagg ctttcttcct cgctgaaagt 1020
gctttacaac ccgaaagcct tcttcacaca cgcggcatgg ctggatcagg ctttcgccca 1080
ttgtccaata ttccccactg ctgcctcccg taggagtctg ggccgtgtct cagtcccagt 1140
gtggctgatc atcctctcag accagctacg gatcgtcgcc ttggtaagcc gttaccttac 1200
caactagcta atccgacata ggctcatccg atagcgcgaa gtccgaagat cctccgcttt 1260
ctcccgtagg acgtatgcgg tattagcgtg ggtttcccca cgttatcccc cactaccggg 1320
cagattccta tgcattactc acccgtccgc cgctcgccac cagggagcaa gctccccgtg 1380
ctgcc 1385
Claims (7)
1. A strain of Zeshu salina fungus with salt-resistant and growth-promoting functions is characterized in that: the strain is named as Zernia zizanioides (Salinicola zeshunii) JP11, and the strain is preserved in China general microbiological culture Collection center (CGMCC) at 9-14 th 2021 with the preservation number of CGMCC No. 23407.
2. A liquid medium suitable for fermentation of the Zygus lucidus halophyte of claim 1, wherein the liquid medium isThe method comprises the following steps: the fermentation medium comprises the following components: 40g L-1Glucose, 10g L-1Yeast powder, 20g L-1Peptone, 80g L-1Sodium chloride, 2g L-1Potassium sulfate, 0.01g L-1Calcium chloride, 1g L-1Potassium feldspar, 2g L-1And (3) lecithin, and adjusting the pH value to 7.5-8.0.
3. The use of the salt-tolerant growth-promoting Zygroshima strain of claim 1 for alleviating the inhibition of salt stress on rice growth.
4. Use according to claim 3, characterized in that: the salt stress refers to the condition of farmland soil with the salt concentration mass ratio not less than 3 per thousand in the soil.
5. The application according to claim 3 or 4, characterized in that it relates to a method of:
(1) inoculating the activated zephyranthus salina strain JP11 seed solution into the zephyranthus salina strain fermentation liquid culture medium according to the volume ratio of 2-5%, and culturing at 28-30 ℃ and 180rpm for 48-60h to obtain a strain suspension containing the zephyranthus salina;
(2) soaking rice seeds in tap water for 24 +/-2 hours, taking full seeds with consistent sizes, and uniformly arranging the seeds in a flowerpot filled with equivalent sterilized nutrient soil poured with 7 per mill of NaCl solution; diluting the fermented ZE-SHI salina fungus suspension with different gradients to obtain 1-5 × 107cfu/mL、1～5×108cfu/mL and 1-5X 109cfu/mL three concentrations of the Zymus salina JP11 bacterial suspension; respectively irrigating three kinds of bacteria liquid with the same amount and different concentrations into the flowerpot, and taking the inactivated bacteria liquid as a reference; and pouring 7 per mill NaCl solution into all the flowerpots; irrigating with 7% NaCl solution every other day; observing the growth condition of the rice at the seedling stage after ten days, and detecting the content of soluble potassium and soluble phosphorus in the soil around the roots of the rice; the results show that: the growth vigor of the rice seedlings applying the JP11 bacterial liquid is obviously superior to that of a control group, and the bacterial strain JP11 is proved to be capable of obviously relieving salt stress pairsThe growth of rice is inhibited, and meanwhile, the content of soluble potassium in soil is increased by 25.4 percent, and the content of soluble phosphorus in soil is increased by 45.9 percent.
6. The use of the eupatorium lucidum having salt tolerance and growth promoting effects of claim 1 for increasing the soluble potassium content in soil, wherein the soluble potassium content in soil is increased by up to 25.4%.
7. The use of the eupatorium lucidum having salt tolerance and growth promoting effects of claim 1 for increasing the soluble phosphorus content in soil, wherein the soluble phosphorus content in soil is increased by 45.9%, and the phosphate solubilizing index are 2.4 and 2.7, respectively.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114540233A (en) * | 2022-02-25 | 2022-05-27 | 广东省科学院生态环境与土壤研究所 | Tailing soil sulfur oxidizing bacteria and application thereof |
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| CN109652339A (en) * | 2019-01-28 | 2019-04-19 | 南京工业大学 | One plant of oil degradation bacterial strain and its application |
| CN112094766A (en) * | 2020-06-09 | 2020-12-18 | 浙江万里学院 | Manganese oxidizing bacteria and application thereof |
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2021
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| CN102994428A (en) * | 2012-11-30 | 2013-03-27 | 辽宁省海洋水产科学研究院 | Marine surfactant-producing bacterial strain LHOD-1 and application thereof |
| CN109652339A (en) * | 2019-01-28 | 2019-04-19 | 南京工业大学 | One plant of oil degradation bacterial strain and its application |
| CN112094766A (en) * | 2020-06-09 | 2020-12-18 | 浙江万里学院 | Manganese oxidizing bacteria and application thereof |
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| CAO等: "Salinicola zeshunii sp nov., a Moderately Halophilic Bacterium Isolated from Soil of a Chicken Farm.", 《CURRENT MICROBIOLOGY》 * |
| MESA等: "Endophytic Cultivable Bacteria of the Metal Bioaccumulator Spartina maritima Improve Plant Growth but Not Metal Uptake in Polluted Marshes Soils.", 《FRONTIERS IN MICROBIOLOGY》 * |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114540233A (en) * | 2022-02-25 | 2022-05-27 | 广东省科学院生态环境与土壤研究所 | Tailing soil sulfur oxidizing bacteria and application thereof |
| CN114540233B (en) * | 2022-02-25 | 2023-06-23 | 广东省科学院生态环境与土壤研究所 | Tailing soil sulfur oxidizing bacteria and application thereof |
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| CN113897320B (en) | 2022-07-26 |
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