CN112625981A - Serratia marcescens and application thereof - Google Patents
Serratia marcescens and application thereof Download PDFInfo
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
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- C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
- C09K17/00—Soil-conditioning materials or soil-stabilising materials
- C09K17/14—Soil-conditioning materials or soil-stabilising materials containing organic compounds only
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
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Abstract
The invention discloses a serratia marcescens strain and application thereof. The strain is named as Serratia marcescens (Serratia marcescens subsp. sakuensis) C273, and the preservation number is GDMCC NO: 61042 the strain is deposited in Guangdong province culture collection of microbial strains of building 5 of Tokyo No. 59 of Tokyo No. 100, Tokyo No. 6, 6 and 3, 2020. Experiments show that the serratia marcescens can remove cadmium ions in soil and water in a co-metabolism mode and can also reduce the pH value in a system so as to dissolve insoluble phosphorus and potassium substances, so that the serratia marcescens has the potential of removing the cadmium ions and improving the effective phosphorus and potassium contents in soil, and can be used for environmental pollution control and plant growth promotion.
Description
Technical Field
The invention belongs to the technical field of inorganic pollutant microbial remediation, and particularly relates to serratia marcescens and application thereof.
Background
Cadmium (Cadmium, Cd), one of the heavy metals, is not an essential element for the growth and development of the organism, and has a serious toxic effect after being absorbed and enriched by passive plants and human bodies. Under the original natural condition, the environmental concentration of cadmium is lower than the toxic concentration of animals and plants, the distribution range is limited, cadmium is in a circulating equilibrium state, and the cadmium does not cause pollution and toxic action to the environment. With the development of industry, cadmium and cadmium compounds are increasingly enriched locally, which causes serious pollution and toxicity to the enriched environment and poses serious threat to the ecological environment, animals, plants and human bodies.
Cadmium differs from other organic pollutants by its structural characteristics, it cannot be decomposed and destroyed; cadmium concentration can only be reduced or eliminated by curing or removal, etc., to achieve a mitigation of the toxic effects. The existing cadmium removal methods are mainly classified into three types, chemical methods, physical methods and biological methods. The method is limited due to the defects of large engineering quantity, high cost, long time consumption, serious secondary pollution and the like of physical and chemical methods. The microbial remediation in the biological method is economic and energy-saving, has the characteristics of strong specificity, small secondary pollution and the like, is widely researched and popularized and is widely applied to sewage treatment and soil pollution treatment.
Disclosure of Invention
The invention aims to overcome the defects in the prior art and provide a serratia marcescens strain.
The invention also aims to provide application of the serratia marcescens in removing cadmium ions in soil and/or water.
Still another object of the present invention is to provide the use of said serratia marcescens for solubilizing insoluble phosphorus and/or potassium compounds.
Still another object of the present invention is to provide the use of said serratia marcescens for increasing available phosphorus and/or potassium in soil.
The purpose of the invention is realized by the following technical scheme:
serratia marcescens, namely Serratia marcescens subsp.sakuensis C273, with the collection number GDMCC NO: 61042 the strain is deposited in Guangdong province culture collection of microbial strains of building 5 of Tokyo No. 59 of Tokyo No. 100, Tokyo No. 6, 6 and 3, 2020.
The 16S rDNA sequence of the serratia marcescens consists of 1434 basic groups (bp), and the nucleotide sequence is shown as SEQ ID NO. 1.
A method for culturing the serratia marcescens comprises the following specific steps: inoculating the serratia marcescens into a culture medium, and culturing at 28-37 ℃.
The culture medium is LB culture medium, MSM culture medium, and contains Cd2+The MSM medium of (1), containing Cd2+And peptone, one of MSM medium, PKO medium and potassium bacteria medium; preferably LB medium.
The Cd content2+The MSM medium contains Cd2+Has a concentration of 10 to 80 mg.L-1。
The Cd content2+And Cd in MSM Medium of peptone2+Has a concentration of 10 to 80 mg.L-1The concentration of peptone is 10 g.L-1。
The formula of the PKO culture medium is as follows: glucose 10 g.L-1，(NH4)2SO4 0.5g·L-1，NaCl 0.2g·L-1，MgSO4·7H2O 0.1g·L-1，KCl 0.3g·L-1，MnSO4·4H2O 0.03g·L-1，FeSO40.003g·L-1，Ca3(PO4)2 5.0g·L-10.5 g.L of yeast powder-120.0 g.L of agar powder-1，pH 6.8～7.0。
The potassium bacteria culture medium comprises the following formula: potassium feldspar 2.5 g.L-1，Na2HPO4 0.2g·L-1，MgSO4·7H2O 0.02g·L-1，NaCl 0.2g·L-1，CaCO3 5.0g·L-1，CaSO4·2H2O 0.1g·L-1Glucose 10 g.L-120.0 g.L of agar powder-1And adjusting the pH value to 6.8-7.0.
The temperature of the culture is preferably 28-30 ℃; more preferably 30 deg.c.
The culture time is 18-48 h; preferably 20-48 h.
The cultivation is carried out in a shaking table, and the rotating speed of the shaking table is 125-150 rpm.
The serratia marcescens is applied to the aspect of removing cadmium ions (reducing the cadmium content in the environment) in soil and/or water.
The serratia marcescens is applied to the aspect of removing cadmium ions in soil and/or water, in order to add the serratia marcescens into the soil and/or water environment containing the cadmium ions, the strain can resist the growth of heavy metal cadmium and can eliminate the cadmium ions in the soil or water so as to remove or reduce the effective concentration of the cadmium ions in the environment.
The cadmium ions are divalent cadmium ions (Cd)2+)。
The concentration of cadmium ions in the water body containing the cadmium ions is 10-80 mg.L-1。
The removal time is more than 6d (days).
The serratia marcescens is applied to the aspect of dissolving insoluble phosphorus and/or potassium compounds.
The application of the serratia marcescens in dissolving insoluble phosphorus and/or potassium compounds is to inoculate the serratia marcescens into a culture medium containing insoluble phosphorus and/or potassium compounds and culture the serratia marcescens at the temperature of 28-37 ℃, wherein the serratia marcescens can reduce the pH value of a system to dissolve the insoluble phosphorus and/or potassium compounds.
The insoluble phosphorus compound is preferably tricalcium phosphate (Ca)3(PO4)2)。
The insoluble potassium compound comprises a potassium-containing mineral; preferably potassium feldspar (K)2O·Al2O3·6SiO2)。
The culture medium containing the insoluble phosphorus compound is preferably a PKO culture medium, and the formula is as follows: glucose 10 g.L-1，(NH4)2SO4 0.5g·L-1，NaCl 0.2g·L-1，MgSO4·7H2O 0.1g·L-1，KCl 0.3g·L-1，MnSO4·4H2O 0.03g·L-1，FeSO4 0.003g·L-1，Ca3(PO4)2 5.0g·L-10.5 g.L of yeast powder-120.0 g.L of agar powder-1，pH 6.8～7.0。
The culture medium containing the insoluble potassium compound is preferably a potassium bacteria culture medium, and the formula is as follows: potassium feldspar 2.5 g.L-1，Na2HPO4 0.2g·L-1，MgSO4·7H2O 0.02g·L-1，NaCl 0.2g·L-1，CaCO3 5.0g·L-1，CaSO4·2H2O 0.1g·L-1Glucose 10 g.L-120.0 g.L of agar powder-1And adjusting the pH value to 6.8-7.0.
The inoculation amount of the serratia marcescens is 1-5% by volume; preferably 2% by volume.
The temperature of the culture is preferably 28-30 ℃.
The culture time is more than 18 h; preferably 18h to 6 d.
The serratia marcescens is applied to the preparation of the cadmium ion passivator.
The serratia marcescens is applied to increasing available phosphorus and/or available potassium in soil.
The application of the serratia marcescens in increasing the available phosphorus and/or the available potassium in the soil is that the serratia marcescens can be added into the soil, and the serratia marcescens can reduce the pH value of a system so as to dissolve insoluble phosphorus and/or potassium compounds in the soil, so that the content of the available phosphorus and the available potassium in the soil is increased, and the plant growth is promoted.
The insoluble phosphorus compound is preferably tricalcium phosphate (Ca)3(PO4)2)。
The insoluble potassium compound comprises a potassium-containing mineral; preferably potassium feldspar (K)2O·Al2O3·6SiO2)。
A biological agent (cadmium ion passivator) for removing cadmium ions in the environment contains the serratia marcescens.
The environment comprises a soil environment and a water body environment.
A biological agent for solubilizing insoluble phosphorus and/or potassium compounds, which comprises the above Serratia marcescens.
After the serratia marcescens C273 grows on the LB plate for 18-20 h, the bacterial plaque is white, round, irregular in edge, flat and glossy, and the diameter of the bacterial colony is about 1-2 mm after the bacterial colony is cultured for 18h at 37 ℃. Can be used in the concentration of cadmium ions of 10-80 mg.L-1Contains 10 g.L-1Peptone grew well in an inorganic salt liquid medium.
Compared with the prior art, the invention has the following advantages and effects:
(1) the serratia marcescens C273 can remove cadmium ions in a co-metabolism mode, and the strain C273 is inoculated to the strain with the cadmium ion concentration of 10-80 mg.L-1Contains 10 g.L-1In an inorganic salt liquid culture medium of peptone, the removal efficiency after 6d of cadmium ions is 40.08%, 40.74%, 38.71% and 49.81% respectively after shaking culture at 30 ℃ and 150rpm for 6 d. The bacterial strain C273 has the advantages of strong resistance to the growth of heavy metal cadmium and removal of cadmium ions, and can be applied to removal of high-concentration cadmium ions in soil and water.
(2) The serratia marcescens C273 can reduce the pH value in the system (possibly secretes organic acid substances such as oxalic acid, formic acid, acetic acid, citric acid, succinic acid and the like to ensure that the pH value in the system is low), so as to dissolve insoluble phosphorus and potassium substances, the strain C273 is inoculated in a PKO or potassium bacteria culture medium, the constant temperature culture is carried out for 48 hours at 30 ℃, and the qualitative test of phosphorus dissolving and potassium dissolving is carried out: after the strain C273 is spotted on a PKO or potassium bacteria culture medium for 6 days, transparent circles appear around the bacteria, which shows that the strain C273 can be applied to cadmium removal in liquid and soil, has the potential of improving the effective phosphorus content of the soil, and can be used for producing microbial inoculum for promoting plant growth.
Drawings
FIG. 1 is a colony morphology map of strain C273 (dish diameter 9 cm).
FIG. 2 is a phylogenetic tree of strain C273.
FIG. 3 shows that the strain C273 contains 10 g.L-1The effect of peptone on removal of cadmium ions in MSM medium is shown.
FIG. 4 is a qualitative test chart of phosphorus and potassium dissolution of the strain C273 (transparent circles show that the strain C has a phosphorus and potassium dissolution function); wherein, A is a strain C273 phosphorus dissolution qualitative test; and B is a qualitative test of potassium dissolution of the strain C273.
Detailed Description
The present invention will be described in further detail with reference to examples, but the embodiments of the present invention are not limited thereto. Reagents, methods and apparatus used in the present invention are conventional in the art unless otherwise indicated. The test methods in the following examples, in which specific experimental conditions are not specified, are generally performed according to conventional experimental conditions or according to the experimental conditions recommended by the manufacturer. Unless otherwise specified, reagents and starting materials for use in the present invention are commercially available.
EXAMPLE 1 screening, isolation and purification and identification of Strain C273
Screening method of strain C273 resistant to cadmium and capable of removing cadmium ions
1. Material preparation
Strain screening soil source: the weed rhizosphere soil of the paddy field around the Dabaoshan, Shaoguan city, Guangdong province is taken, the soil sample is sealed by a sampling bag, and the soil sample is brought back to a laboratory at 4 ℃ for storage and standby.
LB culture medium: 10.0g of peptone, 5.0g of yeast extract powder, 10.0g of NaCl, 7.0-7.2 of pHs, and fixing the volume to 1L of distilled water; the solid medium was supplemented with 18.0g of agar powder. Sterilizing at 121 deg.C for 15 min.
Inorganic salt medium (MSM medium): 5mL of phosphoric acid buffer solution (KH)2PO4 8.5g·L-1、K2HPO4·H2O 21.75g·L-1、Na2HPO4·12H2O 33.4g·L-1、NH4 Cl 5.0g·L-1)，3.0mL 22.5g·L-1MgSO (2) of4Solution (MgSO)4·7H2O 46.125g·L-1)，1.0mL 0.25g·L-1FeCl of3Solution (FeCl)3·6H2O 0.42g·L-1)，1.0mL 36.4g·L-1In (C) is2Solution (CaCl)2·2H2O 48.22g·L-1) 1.0mL of a solution of trace elements (MnSO)4·H2O 39.9mg·L-1，ZnSO4·H2O 42.8mg·L-1，(NH4)6Mo7O24·4H2O 34.7mg·L-1) Mixing, adjusting the pH to 7.0-7.2, adding pure water to a constant volume of 1L, and sterilizing at 121 ℃ for 15 min.
Containing 10 g.L-1MSM medium for peptone: adding 10.0g of tryptone into 1L of MSM culture medium, performing sterilization at the temperature of 121 ℃ for 15min and at the pH of 7.0-7.2.
2. Laboratory apparatus and device
Vertical pressure steam sterilization pan (BL-50A, Shanghai Silique industries, Ltd.), portable pH meter (PHB-4, Shanghai precision science, Ltd.), Centrifuge (Centrifuge 5810R), electric heating oven (DGG-9070A, Shanghai Senxin experiment apparatus, Ltd.), digital display constant temperature water bath pan (HH series, Changzhou national apparatus manufacturing Co., Ltd.), refrigerator (RCD-205AG7, Hai Xin electric appliance), biochemical incubator (PYX-208S-A, Keli apparatus), clean bench (SW-CJ-1F, Sujing Antai air technology, Ltd.), voro mutex mixer (XW-80A, Shanghai Jing industries, Ltd.), MyCycler PCR (BIO-RAD, USA), electrophoresis apparatus (DYY-6C, Bei six instrument works), NaDrno nucleic acid protein quantitative detector (German rmmo), Germany, Gel imaging system (BIO-RAD, USA), floor type constant temperature oscillator (HZQ-211C).
3. Enrichment screening and separation purification of bacterial strain
(1) Isolation and purification of the strains
Weighing 1g of collected soil, adding into 50mL of soil containing 0.5 mg.L-1Cd and 10 g.L-1Domestication culture of peptone in MSM medium (i.e. CdCl)2Adding into a reactor containing 10 g.L-1Culturing Cd in MSM medium of peptone2+To a final concentration of 0.5 mg.L-1) After acclimatization for two days, the cells were inoculated with 2% (v/v) of inoculum size to 1 mg.L-1Cd2+And 10 g.L-1Acclimatization of peptone in MSM medium, and then transfer to Cd concentrations of 2.5, 5 and 10 mg.L at an inoculum size of 2% (v/v), respectively-1Cd2+Contains 10 g.L-1Acclimatization in MSM medium of peptone. After acclimatization and screening, diluting the bacterial liquid to 10 degrees with a culture medium6、107、108And 109Taking 0.2mL of diluent, respectively, coating the diluent on an LB solid culture medium, and culturing at 30 ℃ until bacterial colonies are formed; then selecting the grown single colony, separating and purifying.
(2) Strain screening
And (2) inoculating the purified strain obtained by separation and purification in the step (1) into an LB liquid culture medium for amplification culture for 20-24 h. After activation, the mixture is added to a mixture containing 10 mg.L according to the proportion of 2% (v/v)-1 Cd 2+10 g.L of-1Culturing in MSM culture medium of peptone at 150rpm and 30 deg.C, and collecting bacterial liquid after 6 d. The bacterial liquid was centrifuged at 8000rpm for 10min, the supernatant was collected and passed through a 0.22 μm filter membrane, and the filtrate was measured by flame Atomic Absorption Spectrometry (AAS) (specific references: Zheng 22531, conception, spring, etc.. Shaoguan mining area, rice soil and rice, and risk evaluation [ J]The science of agricultural environment, 2018,37(5):915-2+Removal rate (final concentration/initial concentration x 100%). Screening to obtain Cd2+The strain with the removal efficiency higher than 30% was named strain C273.
Secondly, observing the morphological characteristics of the bacterial colony
The strain C273 grows faster on an LB culture medium and can grow at the temperature of 30-37 ℃. The bacterial plaque is white, round, irregular in edge, flat and glossy, and the diameter of the bacterial colony is about 1-2 mm after the bacterial colony is cultured for 18 hours at 37 ℃ (figure 1). Can be in Cd2+The growth is good in MSM culture medium with 10g/L peptone and 10-80 mg/L peptone.
2.16S rDNA amplification
And (3) amplifying by using the extracted total DNA of the bacteria as a template and adopting a bacterial 16S rDNA universal primer, wherein the forward primer is 27 f: 5'-AGAGTTTGATCCTGGCTCAG-3', reverse primer 1492 r: 5'-GGTTACCTTGTTACGACTT-3' the 16S rDNA gene sequence was amplified.
The total PCR reaction was 25 μ L: 2 mul of each of the upstream and downstream primers, 0.5 mul of template DNA, 12.5 mul of 2 XTaq PCR Master Mix, sterile ultra pure to 25 mul of total volume.
The PCR reaction program is: pre-denaturation at 95 ℃ for 3min, denaturation at 95 ℃ for 50s, annealing at 56 ℃ for 50s, extension at 72 ℃ for 50s, and 35 cycles; and finally, supplementary extension is carried out for 5min at 72 ℃. After the PCR was completed, the PCR product was detected by 1.0% agarose gel electrophoresis, and DL 2000Marker was selected. And observing under a gel imaging system, wherein a clear bright band appears in the middle range of the Marker 1000bp and 2000bp bands.
3.16 determination of the S rDNA sequence
The PCR amplified product was sequenced by Biotechnology of Boxing Ke, Beijing Rui (Guangzhou division). The 16S rDNA gene sequence of the obtained strain is as follows (SEQ ID NO. 1):
GCTTACCTGCAGTCGAGCGGTAGCACGGGGGAGCTTGCTCCCTGGGTGACGAGCGGCGGACGGGTGAGTAATGTCTGGGAAACTGCCTGATGGAGGGGGATAACTACTGGAAACGGTAGCTAATACCGCATAACGTCGCAAGACCAAAGAGGGGGACCTTCGGGCCTCTTGCCATCAGATGTGCCCAGATGGGATTAGCTAGTAGGTGGGGTAATGGCTCACCTAGGCGACGATCCCTAGCTGGTCTGAGAGGATGACCAGCCACACTGGAACTGAGACACGGTCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGCACAATGGGCGCAAGCCTGATGCAGCCATGCCGCGTGTGTGAAGAAGGCCTTCGGGTTGTAAAGCACTTTCAGCGAGGAGGAAGGTGGTGAGCTTAATACGCTCATCAATTGACGTTACTCGCAGAAGAAGCACCGGCTAACTCCGTGCCAGCAGCCGCGGTAATACGGAGGGTGCAAGCGTTAATCGGAATTACTGGGCGTAAAGCGCACGCAGGCGGTTTGTTAAGTCAGATGTGAAATCCCCGGGCTCAACCTGGGAACTGCATTTGAAACTGGCAAGCTAGAGTCTCGTAGAGGGGGGTAGAATTCCAGGTGTAGCGGTGAAATGCGTAGAGATCTGGAGGAATACCGGTGGCGAAGGCGGCCCCCTGGACGAAGACTGACGCTCAGGGGCGAAAGCGTGGGGAGCAAACAGGATTAGATACCCCTGGTAGTCCACGCTGTAAACGATGTCGATTTTGGAGGTTGTGCCCCTTGAGGCGTGGCTTCCGGAAGCTAACGCGTTAAATCGACCGCCTGGGGAGTACGGCCGCAAGGTTAAAACTCAAATGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGATGCAACGCGAAGAACCTTACCTACTCTTGACATCCAGAGAACTTAGCAGAGATGCTTTGGTGCCTTCGGGAACTCTGAGACAGGTGCTGCATGGCTGTCGTCAGCTCGTGTTGTGAAATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTATCCTTTGTTGCCAGCGGTTCGGCCGGGAACTCAAAGGAGACTGCCAGTGATAAACTGGAGGAAGGTGGGGATGACGTCAAGTCATCATGGCCCTTACGAGTAGGGCTACACACGTGCTACAATGGCGTATACAAAGAGAAGCGACCTCGCGAGAGCAAGCGGACCTCATAAAGTACGTCGTAGTCCGGATTGGAGTCTGCAACTCGACTCCATGAAGTCGGAATCGCTAGTAATCGTAGATCAGAATGCTACGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGGGAGTGGGTTGCAAAAGAAGTAGGTAGCTTAACCTTCGGGAGGGCGCTTACCACTTGGATCATT。
the above sequence consists of 1434 bases (bp).
The obtained 16S rDNA gene sequence is submitted to a National Center for Biological Information (NCBI) webpage for BLAST comparison, and is subjected to homology comparison analysis with the 16S rDNA gene of a relevant model strain in an LPSN database (http:// www.bacterio.net/index. html), a model strain sequence with higher homology is downloaded, and an amplification product sequence is subjected to BLAST comparison and homology analysis on a National Center for Biological Information (NCBI) website, and a system development tree is constructed by a Neightbour-Joining method by adopting Mega 6.0 software. Comparison of the 16S rDNA sequence revealed that the strain C273 had 99.09% homology with Serratia marcescens subsp.sakuensis KREDT (FIG. 2).
Fourthly, identifying the strain C273 as a new functional strain
According to the colony morphological characteristics and the molecular biological identification result of the strain C273, the strain C273 is named as Serratia marcescens subsp. The strain is preserved in Guangdong province microorganism culture collection center (GDMCC) with the preservation number of GDMCC NO: 61042, 6 months and 3 days in 2020. The address of the preservation unit is No. 59 building No. 5 building of No. 100 college of Jifura Zhonglu, Guangzhou city.
At present, the number of the current day,at home and abroad, no document reports that Serratia marcescens (Serratia marcocens subsp. sakuensis) has the functions of resisting the growth of heavy metal cadmium and removing cadmium. Therefore, the serratia marcescens C273 strain can reduce Cd in solution2+Novel strains of function.
Example 2 Strain C273 in the presence of Cd2+Cd in inorganic salt culture medium of peptone2+Removal of
Streaking the strain C273, culturing at 30 deg.C for 20 hr on LB plate, selecting single colony, inoculating to LB liquid culture medium, culturing at 30 deg.C and 150rpm in shaking table for 20 hr, and inoculating to Cd at a ratio of 2% (v/v)2+Concentrations of 10, 20, 40 and 80 mg.L-1Containing 10 g.L-1Culturing tryptone in MSM medium, and sampling after 6d (days); 3 replicates were set up with no addition of strain S1 as a Control (CK). Centrifuging the sample solution at 80000rpm for 10min, filtering the supernatant with 0.22 μm filter membrane, and measuring Cd by flame Atomic Absorption Spectrometry (AAS)2+The same procedure as in example 1.
Cd after 6 days of culture2+The removal efficiency is shown in fig. 3: the results show that Cd is obtained after adding the strain C2732+The concentration decreased significantly (blank removal was 0 and therefore not shown in the figure). Cd [ Cd ]2+Initial concentrations were 10, 20, 40 and 80 mg.L-1Then, Cd2+The removal efficiencies were 40.08%, 40.74%, 38.71%, 49.81%, respectively.
EXAMPLE 3 qualitative test of phosphorus and Potassium solubilization of Strain C273
Phosphorus and potassium solubilizing bacteria (Phosphate/potassium solubilizing bacteria) of strain C273, meaning the conversion of insoluble phosphorus and potassium elements into soluble phosphorus and potassium elements (specific references: Rawat, P., Das, S., Shankhdhar, D., et al., Phosphate-solubilizing microorganisms: Mechanism and the role in Phosphate solubilization and uptake [ J ]. Journal of Soil Science and Plant nutrition.2020, https:// doi.org/10.1007/s 42729-020-: inoculating a strain C273 marked on an LB solid culture medium into an LB liquid culture medium, shaking and culturing for 18-24 h at 125-150 rpm and 28-30 ℃ in a shaking table, then respectively dotting the strain C on a PKO culture medium and a potassium bacteria solid culture medium, inoculating 2% of the strain C on the PKO liquid culture medium and the potassium bacteria liquid culture medium, setting 3 repeated tests, placing the strain C in a 30 ℃ incubator or the shaking table (125-150 rpm), setting 3 repeated tests, placing the strain C in a 30 ℃ incubator or the shaking table 6, observing whether a transparent ring formed after insoluble phosphorus compounds and potassium minerals are dissolved is arranged around a colony in the culture medium, observing the size of the transparent ring, and taking a picture. Whether the strain has the effect of dissolving the indissolvable phosphorus compounds and the potassium minerals is judged according to the hydrolysis ring. Wherein the content of the first and second substances,
PKO culture medium formula: glucose 10 g.L-1，(NH4)2SO4 0.5g·L-1，NaCl 0.2g·L-1，MgSO4·7H2O 0.1g·L-1，KCl 0.3g·L-1，MnSO4·4H2O 0.03g·L-1，FeSO4 0.003g·L-1，Ca3(PO4)25.0g·L-10.5 g.L of yeast powder-120.0 g.L of agar powder-1pH is 6.8-7.0 (no agar powder is added into a liquid culture medium);
potassium bacteria culture medium: potassium feldspar 2.5 g.L-1，Na2HPO4 0.2g·L-1，MgSO4·7H2O 0.02g·L-1，NaCl 0.2g·L-1，CaCO3 5.0g·L-1，CaSO4·2H2O 0.1g·L-1Glucose 10 g.L-120.0 g.L of agar powder-1And adjusting the pH value to 6.8-7.0 (agar powder is not added in the liquid culture medium).
The results are shown in FIG. 4: strain C273 shows hydrolysis loop on PKO medium and potassium-dissolving bacteria solid medium, and shows that it has dissolved insoluble phosphorus compound (Ca)3(PO4)2) Or potassium mineral (potassium feldspar), and the pH of the solution is reduced to about 4.3.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Sequence listing
<110> southern China university of agriculture
<120> serratia marcescens and application thereof
<160> 3
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1434
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<220>
<223> 16S rDNA gene sequence
<400> 1
gcttacctgc agtcgagcgg tagcacgggg gagcttgctc cctgggtgac gagcggcgga 60
cgggtgagta atgtctggga aactgcctga tggaggggga taactactgg aaacggtagc 120
taataccgca taacgtcgca agaccaaaga gggggacctt cgggcctctt gccatcagat 180
gtgcccagat gggattagct agtaggtggg gtaatggctc acctaggcga cgatccctag 240
ctggtctgag aggatgacca gccacactgg aactgagaca cggtccagac tcctacggga 300
ggcagcagtg gggaatattg cacaatgggc gcaagcctga tgcagccatg ccgcgtgtgt 360
gaagaaggcc ttcgggttgt aaagcacttt cagcgaggag gaaggtggtg agcttaatac 420
gctcatcaat tgacgttact cgcagaagaa gcaccggcta actccgtgcc agcagccgcg 480
gtaatacgga gggtgcaagc gttaatcgga attactgggc gtaaagcgca cgcaggcggt 540
ttgttaagtc agatgtgaaa tccccgggct caacctggga actgcatttg aaactggcaa 600
gctagagtct cgtagagggg ggtagaattc caggtgtagc ggtgaaatgc gtagagatct 660
ggaggaatac cggtggcgaa ggcggccccc tggacgaaga ctgacgctca ggggcgaaag 720
cgtggggagc aaacaggatt agatacccct ggtagtccac gctgtaaacg atgtcgattt 780
tggaggttgt gccccttgag gcgtggcttc cggaagctaa cgcgttaaat cgaccgcctg 840
gggagtacgg ccgcaaggtt aaaactcaaa tgaattgacg ggggcccgca caagcggtgg 900
agcatgtggt ttaattcgat gcaacgcgaa gaaccttacc tactcttgac atccagagaa 960
cttagcagag atgctttggt gccttcggga actctgagac aggtgctgca tggctgtcgt 1020
cagctcgtgt tgtgaaatgt tgggttaagt cccgcaacga gcgcaaccct tatcctttgt 1080
tgccagcggt tcggccggga actcaaagga gactgccagt gataaactgg aggaaggtgg 1140
ggatgacgtc aagtcatcat ggcccttacg agtagggcta cacacgtgct acaatggcgt 1200
atacaaagag aagcgacctc gcgagagcaa gcggacctca taaagtacgt cgtagtccgg 1260
attggagtct gcaactcgac tccatgaagt cggaatcgct agtaatcgta gatcagaatg 1320
ctacggtgaa tacgttcccg ggccttgtac acaccgcccg tcacaccatg ggagtgggtt 1380
gcaaaagaag taggtagctt aaccttcggg agggcgctta ccacttggat catt 1434
<210> 2
<211> 20
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<220>
<223> the forward primer was 27f
<400> 2
<210> 3
<211> 19
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<220>
<223> reverse primer is 1492r
<400> 3
ggttaccttg ttacgactt 19
Claims (10)
1. A serratia marcescens strain is characterized in that: the name is Serratia marcocens subsp.sakuensis C273, the collection number is GDMCC NO: 61042 the strain is deposited in Guangdong province culture collection of microbial strains of building 5 of Tokyo No. 59 of Tokyo No. 100, Tokyo No. 6, 6 and 3, 2020.
2. A method for culturing Serratia marcescens according to claim 1, which comprises the following steps: inoculating the serratia marcescens into a culture medium, and culturing at 28-37 ℃.
3. Use according to claim 2, characterized in that:
the culture medium is LB culture medium, MSM culture medium, and contains Cd2+The MSM medium of (1), containing Cd2+And peptone, one of MSM medium, PKO medium and potassium bacteria medium;
the Cd content2+The MSM medium contains Cd2+Has a concentration of 10 to 80 mg.L-1；
The Cd content2+And Cd in MSM Medium of peptone2+Has a concentration of 10 to 80 mg.L-1The concentration of peptone is 10 g.L-1；
The culture time is 18-48 h.
4. Use of serratia marcescens according to claim 1 for removing cadmium ions from soils and/or water bodies.
5. Use according to claim 4, characterized in that: adding serratia marcescens into soil and/or water environment containing cadmium ions to remove or reduce the effective concentration of the cadmium ions in the environment;
the cadmium ions are divalent cadmium ions.
6. Use of serratia marcescens according to claim 1 for preparing a cadmium ion inactivator.
7. Use of serratia marcescens according to claim 1 for solubilizing insoluble phosphorus and/or potassium compounds.
8. Use according to claim 7, characterized in that: inoculating serratia marcescens into a culture medium containing insoluble phosphorus and/or potassium compounds, and culturing at 28-37 ℃, wherein the serratia marcescens can dissolve the insoluble phosphorus and/or potassium compounds by low system pH;
the culture time is more than 18 h.
9. Use of serratia marcescens according to claim 1 for increasing available phosphorus and/or potassium in soil.
10. A biological agent for removing cadmium ions in the environment and dissolving insoluble phosphorus and/or potassium compounds, which is characterized in that: comprising the serratia marcescens of claim 1;
the environment is a soil environment and/or a water body environment.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112852688A (en) * | 2021-04-15 | 2021-05-28 | 河南科技学院 | Serratia LE and application thereof |
| CN114134063A (en) * | 2021-08-31 | 2022-03-04 | 华中农业大学 | Strain Serratia sp.X10 for reducing cadmium accumulation in rice and application thereof |
| CN115418326A (en) * | 2022-03-28 | 2022-12-02 | 湖南省蔬菜研究所 | Complex microbial inoculant and application thereof |
| CN114657102B (en) * | 2022-04-07 | 2023-07-14 | 西安建筑科技大学 | Serratia marcescens and application thereof in degradation of tetrabromobisphenol A |
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| CN104560796A (en) * | 2014-12-18 | 2015-04-29 | 湖北工业大学 | Tolerant serratia grimesii for enriching heavy metals and application of serratia grimesii |
| CN110157643A (en) * | 2019-05-23 | 2019-08-23 | 安徽农业大学 | The bacterium H4C8 of cadmium content and its application in a kind of reduction wheat |
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| CN104560796A (en) * | 2014-12-18 | 2015-04-29 | 湖北工业大学 | Tolerant serratia grimesii for enriching heavy metals and application of serratia grimesii |
| CN110157643A (en) * | 2019-05-23 | 2019-08-23 | 安徽农业大学 | The bacterium H4C8 of cadmium content and its application in a kind of reduction wheat |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112852688A (en) * | 2021-04-15 | 2021-05-28 | 河南科技学院 | Serratia LE and application thereof |
| CN114134063A (en) * | 2021-08-31 | 2022-03-04 | 华中农业大学 | Strain Serratia sp.X10 for reducing cadmium accumulation in rice and application thereof |
| CN115418326A (en) * | 2022-03-28 | 2022-12-02 | 湖南省蔬菜研究所 | Complex microbial inoculant and application thereof |
| CN114657102B (en) * | 2022-04-07 | 2023-07-14 | 西安建筑科技大学 | Serratia marcescens and application thereof in degradation of tetrabromobisphenol A |
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| CN112625981B (en) | 2022-03-25 |
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