CN102604832A - Screening method of aureomycin degrading strains - Google Patents
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- CN102604832A CN102604832A CN2012100312540A CN201210031254A CN102604832A CN 102604832 A CN102604832 A CN 102604832A CN 2012100312540 A CN2012100312540 A CN 2012100312540A CN 201210031254 A CN201210031254 A CN 201210031254A CN 102604832 A CN102604832 A CN 102604832A
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Abstract
The invention relates to a screening method of aureomycin degrading strains, and belongs to the technical field of degrading antibiotic residues by microorganisms. The screening method comprises the following particular steps: isolating the aureomycin degrading strains; purifying the strains; verifying the degrading functions of the strains; and determining the degrading activities of the strains. The screening method adopted by the invention is quick, feasible, economic and low-priced, and is suitable to obtain strains capable of degrading aureomycin residues of different concentrations, especially the efficient strains capable of degrading low-concentration or oligotrophic-grade aureomycin. The obtained strains can be used for preparing an aureomycin degrading agent. According to the invention, the aureomycin-producing microbe culture residue and waste water are utilized as the original materials for screening; and the obtained strains and the prepared agents can be directly applied in treating waste of aureomycin production enterprises, and can be possibly applied in remediation of peripheral soil and water environment polluted by aureomycin. According to the invention, high performance liquid chromatography is used for determining the degrading activities of the strains, and the obtained results have high accuracy and precision.
Description
Technical field
The present invention relates to a kind of screening method of duomycin degradation bacteria strains, belong to microbiological deterioration antibiotic remains technical field.
Background technology
Duomycin (Chlortetracycline; Aureomycin), claim Uromycin again, belong to TCs.Chemical name is 6-methyl-4-(dimethylamino)-3,6,10,12,12a-penta hydroxy group-1, and 11-dioxo-7-chloro-1,4,4a, 5,5a, 6,11,12a-octahydro-2-tetracene methane amide, molecular formula is C
22H
23ClN
2O
8, molecular weight is 478.88, chemical structural formula is:
Duomycin is a kind of broad-spectrum antibiotics, all can produce restraining effect to gram-positive microorganism, Gram-negative bacteria, spirochete, Rickettsiae, mycoplasma, chlamydozoan etc., has purposes widely in fields such as medicine, livestock industry.
China is duomycin big producing country, in the duomycin production process, can produce a large amount of bacterium slags and waste water.The bacterium slag is classified as dangerous solid waste because of problem such as duomycin is residual by country; Duomycin in the waste water is residual because of unrestriction emission standard still at present, and duomycin is residual in the waste water might be discharged in the environment, causes environmental pollution, even human health and ecological security are constituted a threat to.
At present, duomycin environment residue problem has become one of focus environmental problem of domestic and international common concern.So far, report few about the research of duomycin biological degradation and degradation bacteria strains screening aspect both at home and abroad, only retrieve the document of 1 piece of relevant degradation bacteria strains screening aspect.What fine jade utilized domestication of duomycin waste water and screening to obtain a strain monascus B3 in 2005, and this bacterial strain is 4000mg/L to the highest tolerance concentration of duomycin, is being under the sole carbon source condition with duomycin, and its degradation rate only has about 10%; Under other carbon source of interpolation and optimum condition, cultivate, its degradation rate can reach 51.5%.The said screening method complex operation of the document, the cycle is long, needs elder generation through after more than 60 days the domestication, carries out bacterial strain screening again; Simultaneously, this method lays particular emphasis on the screening of tolerance higher concentration duomycin degradation bacteria strains, does not consider the screening problem of low-concentration gold mycin degradation bacteria strains.
Obviously; Research and propose a kind of fast, efficient, and the screening method of the different concns duomycin bacterial strain that can obtain simultaneously to degrade; Particularly the degrade efficient bacterial strain of lower concentration or poor trophic level duomycin; To obtain degradation effect farthest, in the research of microbiotic biological degradation theory and method, have important scientific meaning and application prospect.
Summary of the invention
The objective of the invention is for provide a kind of fast, efficient, and the screening method of the different concns duomycin degradation bacteria strains that can obtain simultaneously to degrade.
The objective of the invention is to realize through following technical scheme.
The screening method of a kind of duomycin degradation bacteria strains of the present invention, concrete steps are following:
1) separation of duomycin degradation bacteria strains
Gather duomycin bacterium slag and waste water, then the bacterium slag is joined in the zero(ppm) water, mix, extract bacterium slag leach liquor; The adding proportion of bacterium slag and zero(ppm) water is 1g: 10ml;
Draw 200-400 μ l waste water and 200-400 μ l bacterium slag leach liquor respectively, coat successively on the different inorganic salt solid medium of duomycin content, place under 28 ℃ of-30 ℃ of conditions shading to cultivate 2-7 days, obtain different bacterium colonies; Duomycin content is 0.1g/L-15.0g/L in the substratum;
2) bacterial strain purifying
Step 1) is separated the bacterium colony obtain, rule containing on the inorganic salt solid medium flat board of duomycin respectively, further separate and purifying; This step can repeat 1-2 time, until obtaining single bacterium colony; Then single bacterium colony is inserted in the inorganic salt liquid substratum that contains duomycin successively, place 28 ℃-30 ℃, under the 160-170r/min oscillating condition, shading was cultivated 1-3 days, obtained single bacterium colony nutrient solution;
3) strains for degrading functional verification
With step 2) nutrient solution that obtains, be to rule on the different inorganic salt solid medium flat board of sole carbon source and chlortetracycline concentration at duomycin respectively, 28 ℃ of-30 ℃ of lucifuges were cultivated 2-7 days; It can be the degradation bacteria strains of sole carbon source with duomycin that the bacterial strain that can grow is; Can obtain the chlortetracycline concentration scope that each bacterial strain can utilize and degrade through observing;
4) strains for degrading determination of activity
What step 3) was obtained can be the bacterial strain of sole carbon source growth with duomycin, is transferred to respectively and carries out activation on the activation medium; The inoculum size of then activatory bacterium liquid being pressed 1%-3%; Be transferred to duomycin is in the suitable inorganic salt liquid substratum of sole carbon source and concentration; 28 ℃-30 ℃, the 160-170r/min concussion is dark cultivated 1-3 days, at last with the degradation rate of each bacterial strain of high effective liquid chromatography for measuring to duomycin; The high bacterial strain of different concns degradation rate, 4 ℃ of storages are preserved in the test tube slant.
Beneficial effect
The screening method that the present invention adopts is prone to row fast, and relatively cheap, resulting bacterial strain can be used for preparing the duomycin degradation bacterial agent; The present invention utilizes duomycin bacterium slag and the initial raw material of waste water for screening, and resulting bacterial strain and the microbial inoculum of processing can directly apply to the processing of duomycin enterprise waste, also might be applied to periphery by the reparation of duomycin Contaminated soil and water body environment; The present invention is active with the high effective liquid chromatography for measuring strains for degrading, and gained result's accuracy and precision are higher.
Description of drawings
Fig. 1 is a duomycin all wave band scintigram;
Fig. 2 is a duomycin performance liquid chromatography canonical plotting.
Embodiment
Embodiment 1
1) bacterium sample collecting and pre-treatment
Gather the bacterium slag and the waste water appearance of duomycin manufacturing enterprise, and take by weighing bacterium slag 10g, place the 250mL triangular flask, add 100ml zero(ppm) water, on shaking table, shake up, leach liquor is subsequent use.
2) strains separation
Draw 400 μ l waste water and bacterium slag leach liquor, separate application places under 28 ℃ of-30 ℃ of conditions shading to cultivate 2-7 days on inorganic salt solid medium flat board.The inorganic salt solid medium consists of: (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA 0.015g, glucose 2.0g, agar 13.0g.Above-mentioned substance is dissolved in the 1000ml zero(ppm) water 115 ℃ of moist heat sterilization 15-20min.Falling before the flat board, in substratum, add duomycin, add concentration and be followed successively by 0.1g/L, 1.0g/L, 2.0g/L, 3.0g/L, 4.0g/L, 5.0g/L, 10.0g/L and 15.0g/L, shake up.
3) bacterial strain purifying
Make even and separate the single bacterium colony that obtains on the plate, rule containing on the inorganic salt solid medium of duomycin, further separate and purifying.This step repeats 1-2 time, until obtaining single bacterium colony.Then single bacterium colony is inserted in the inorganic salt liquid substratum that contains duomycin successively, 28 ℃-30 ℃, 170r/min vibration shading was cultivated 1-3 days.
The inorganic salt solid medium consists of: (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA 0.015g, glucose 5.0g/L, agar 13g/L.Above-mentioned substance is dissolved in the 1000ml zero(ppm) water 115 ℃ of moist heat sterilization 15-20min.Fall before the flat board, add 2.0g/L duomycin, shake up.
The inorganic salt liquid substratum consists of: (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA 0.015g, glucose 5.0g/L.Above-mentioned substance is dissolved in the 1000ml zero(ppm) water, 115 ℃ of moist heat sterilization 15-20min, to be cooled after room temperature, add 2.0g/L duomycin, shake up.
4) strains for degrading functional verification
At first, the bacterial strain that purifying is obtained carries out activation culture, bacterium in this way, and with beef-protein medium activation culture 1 day, fungi in this way with Ma Dingshi substratum activation culture 2 days, obtained strain cultured solution.Get nutrient solution then, respectively being to rule on the different inorganic salt solid medium flat board of sole carbon source and chlortetracycline concentration with duomycin, under 28 ℃ of-30 ℃ of conditions, lucifuge was cultivated 2-7 days.It can be the degradation bacteria strains of sole carbon source with duomycin that the bacterial strain that can grow is.Observe each bacterial strain upgrowth situation on different concns duomycin flat board, can obtain the concentration and the scope of the suitable degraded duomycin of each bacterial strain.Part bacterial strain experimental result is seen table 1.
The inorganic salt solid medium consists of: (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl0.2g, CaCl
20.1g, FeSO
40.01g, EDTA 0.015g, agar 13g.Above-mentioned substance is dissolved in the 1000ml zero(ppm) water 121 ℃ of moist heat sterilization 20min.Falling before the flat board, adding duomycin respectively is 0.1g/L, 5.0g/L, 10.0g/L, 15.0g/L, is made into the substratum of 4 duomycin gradients, shakes up.
The beef-protein medium prescription is: Carnis Bovis seu Bubali cream 3.0g, peptone 10.0g, NaCl 5.0g, agar 13.0g.It is dissolved in the 1000mL zero(ppm) water 121 ℃ of moist heat sterilization 20min.
The Ma Dingshi substratum consists of: glucose 10.0g, peptone 5.0g, KH
2PO
40.5g, MgSO
47H
2O 0.5g.It is dissolved in the 1000mL zero(ppm) water 115 ℃ of moist heat sterilization 15-20min.
Strain growth situation under the different chlortetracycline concentrations of table 1
Annotate :+representative growth; ++ represent raised growth;-representative is growth not.
Can know by table 1, the concentration range of duomycin lower concentration (≤0.1g/L), middle lower concentration (0.2-4.9g/L), middle and high concentration (5.0-9.9g/L) and high density (>=10g/L) time, all can obtain degradation bacteria strains;
5) strains for degrading activity identification
The bacterial strain that will have degraded duomycin is transferred to respectively and carries out activation on the inorganic salt liquid substratum; And activatory bacterium liquid is transferred to duomycin by 2% inoculum size is in the suitable inorganic salt liquid substratum of sole carbon source and concentration; Be numbered the LJ236 bacterial strain such as what obtained by step 4), suitable chlortetracycline concentration is 0.1g/L, 28 ℃-30 ℃; The 170r/min concussion is dark cultivated 1-3 days, adopted the degradation rate of each bacterial strain of high effective liquid chromatography for measuring to duomycin then.The high bacterial strain of different concns degradation rate, 4 ℃ of preservations are preserved in the test tube slant.
Duomycin solution all wave band scanning experiment
Get duomycin reference liquid 2mL, join in the tube comparison tubes, be diluted to 25mL.On ultraviolet-visible spectrophotometer, carry out full spectral scan, finding has absorption peak near 280nm and the 375nm, sees accompanying drawing 1.Therefore, duomycin is measured wavelength and can be adopted 280nm and 375nm.
The drafting of typical curve
Accurately take by weighing the duomycin standard specimen; Using volume ratio is that 1: 5 the ultrapure water and the mixing solutions of methyl alcohol dissolve; Be made into duomycin content and be followed successively by the series standard solution of 0.1g/L, 0.2g/L, 0.3g/L, 0.4g/L, 0.5g/L, 0.6g/L; Membrane filtration with 22 μ m obtains filtrating, measures with high performance liquid chromatograph, and the equation of linear regression that obtains chlortetracycline concentration x and peak area y is y=956.5X-11.434.Referring to accompanying drawing 2.
Chromatographic condition: weighting agent is an octadecylsilane chemically bonded silica; Moving phase is oxalic acid solution (0.01mol/L): acetonitrile: the methyl alcohol volume ratio is 10: 3: 2; Flow velocity is 1.0ml/min; Column temperature is a room temperature; Sample size is 20ul; Detect wavelength 375nm.
Machine testing on the sample obtains accurate degradation rate
Get liquid 1mL to be measured, add the methyl alcohol of 5mL, shake up, the centrifugal 5min of 8000r/min gets supernatant, obtains filtrating with the membrane filtration of 22 μ m, measures with high performance liquid chromatograph then, calculates the content and the degradation rate of duomycin according to equation of linear regression.For example, the efficient duomycin degradation bacteria strains LJ220 that screening obtains, when being sole carbon source with duomycin, its degradation rate can reach 31.60%.What introduce in this result and He Yuzai " characteristic research of monascus B3 degraded duomycin waste water " (University of Fuzhou's master thesis, 2005) document is that the degradation results 9.35% of sole carbon source is compared with duomycin, improves a lot.
Embodiment 2
The enrichment of duomycin degradation bacteria strains, isolation and purification
Compare with embodiment 1, this method is carried out the bacterial classification enrichment earlier, carries out isolation and purification again.Comprise:
1) bacterial classification enrichment
Gather bacterium slag and waste water, take by weighing waste residue 10g, place the 250mL triangular flask, add 100ml zero(ppm) water, on shaking table, shake up, extract leach liquor.Draw 10mL waste water appearance and bacterium slag leach liquor, insert in the triangular flask of the 250mL that the 100ml enrichment medium is housed respectively, 28 ℃-30 ℃, under the 170r/min concussion condition, secretly cultivated 2-3 days.Get the 5ml nutrient solution then and change in the new identical substratum, under identical culture condition, continue enrichment culture once.
The inorganic salt liquid enrichment medium consists of: glucose 2g, (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA0.015g, agar 13g.Above-mentioned substance is dissolved in the 1000ml zero(ppm) water; Divide and install in the 250mL triangular flask; 115 ℃ of moist heat sterilization 15-20min; In Bechtop, add duomycin, add concentration and be followed successively by 0.1g/L, 1.0g/L, 2.0g/L, 3.0g/L, 4.0g/L, 5.0g/L, 10.0g/L and 15.0g/L, shake up.
2) strains separation
Draw the above-mentioned enrichment culture liquid of 200 μ l, corresponding one by one respectively coating on the inorganic salt solid medium flat board of top said 8 kinds of different chlortetracycline concentrations places under 28 ℃ of-30 ℃ of conditions shading to cultivate 2-7 days.
The inorganic salt solid medium consists of: glucose 2.0g, (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA0.015g, agar 13.0g is dissolved in the 1000ml zero(ppm) water.115 ℃ of moist heat sterilization 15-20min.Falling before the flat board, in substratum, add duomycin, it is identical with the inorganic salt liquid enrichment medium to add concentration.
Operation steps afterwards is with embodiment 1.
Claims (10)
1. the screening method of a duomycin degradation bacteria strains is characterized in that concrete steps are following:
1) separation of duomycin degradation bacteria strains
Gather duomycin bacterium slag and waste water, the bacterium slag is joined in the zero(ppm) water, mix, extract bacterium slag leach liquor;
Draw waste water and bacterium slag leach liquor respectively, coat successively on the different inorganic salt solid medium flat board of duomycin content, place under 28 ℃ of-30 ℃ of conditions shading to cultivate 2-7 days, obtain different bacterium colonies;
2) bacterial strain purifying
Step 1) is separated the bacterium colony obtain, rule containing on the inorganic salt solid medium flat board of duomycin respectively, further separate and purifying; This step repeats 1-2 time, until obtaining single bacterium colony; Then single bacterium colony is inserted in the inorganic salt liquid substratum that contains duomycin successively, place 28 ℃-30 ℃, under the 160-170r/min oscillating condition, shading was cultivated 1-3 days, obtained single bacterium colony nutrient solution;
3) strains for degrading functional verification
With step 2) nutrient solution that obtains, be to rule on the different inorganic salt solid medium flat board of sole carbon source and chlortetracycline concentration at duomycin respectively, 28 ℃ of-30 ℃ of lucifuges were cultivated 2-7 days; It can be the degradation bacteria strains of sole carbon source with duomycin that the bacterial strain that can grow is;
4) strains for degrading determination of activity
What step 3) was obtained can be the bacterial strain of sole carbon source growth at duomycin, is transferred to respectively and carries out activation on the activation medium; The inoculum size of then activatory bacterium liquid being pressed 1%-3%; Be transferred to duomycin is in the suitable inorganic salt liquid substratum of sole carbon source and concentration; 28 ℃-30 ℃, the 160-170r/min concussion is dark cultivated 1-3 days, at last with the degradation rate of each bacterial strain of high effective liquid chromatography for measuring to duomycin; The high bacterial strain of different concns degradation rate, 4 ℃ of storages are preserved in the test tube slant.
2. the screening method of a kind of duomycin degradation bacteria strains according to claim 1 is characterized in that: the adding proportion of bacterium slag and zero(ppm) water is 1g: 10ml in the step 1), and the volume of drawing waste water and bacterium slag leach liquor is 200-400 μ l.
3. the screening method of a kind of duomycin degradation bacteria strains according to claim 1 is characterized in that: in the step 1) in the substratum duomycin content be 1.0g/L-15.0g/L.
4. the screening method of a kind of duomycin degradation bacteria strains according to claim 1 is characterized in that: the consisting of of inorganic salt solid medium in the step 1): (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA 0.015g, glucose 2g, agar 13g; Above-mentioned substance is dissolved in the 1000ml zero(ppm) water 115 ℃ of moist heat sterilization 15-20min; Falling before the flat board, in substratum, add duomycin, add concentration and be followed successively by 0.1g/L, 1.0g/L, 2.0g/L, 3.0g/L, 4.0g/L, 5.0g/L, 10.0g/L and 15.0g/L, shake up.
5. the screening method of a kind of duomycin degradation bacteria strains according to claim 1 is characterized in that: step 2) in the consisting of of inorganic salt solid medium: (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA 0.015g, glucose 5.0g/L, agar 13g/L; Above-mentioned substance is dissolved in the 1000ml zero(ppm) water 115 ℃ of moist heat sterilization 15-20min; Fall before the flat board, add 2.0g/L duomycin, shake up.
6. the screening method of a kind of duomycin degradation bacteria strains according to claim 1 is characterized in that: step 2) in the inorganic salt liquid substratum consist of: (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA 0.015g, glucose 5.0g/L; Above-mentioned substance is dissolved in the 1000ml zero(ppm) water, 115 ℃ of moist heat sterilization 15-20min, to be cooled after room temperature, add 2.0g/L duomycin, shake up.
7. the screening method of a kind of duomycin degradation bacteria strains according to claim 1 is characterized in that: the inorganic salt solid medium consists of in the step 3): (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl0.2g, CaCl
20.1g, FeSO
40.01g, EDTA 0.015g, agar 13g; Above-mentioned substance is dissolved in the 1000ml zero(ppm) water 121 ℃ of moist heat sterilization 20min; Falling before the flat board, adding duomycin respectively is 0.1g/L, 5.0g/L, 10.0g/L, 15.0g/L, is made into the substratum of 4 duomycin gradients, shakes up.
8. the screening method of a kind of duomycin degradation bacteria strains according to claim 1 is characterized in that: the bacterium activation medium is a beef-protein medium in the step 4), and its prescription is: Carnis Bovis seu Bubali cream 3.0g, peptone 10.0g, NaCl 5.0g, agar 13.0g; It is dissolved in the 1000mL zero(ppm) water 121 ℃ of moist heat sterilization 20min.
9. the screening method of a kind of duomycin degradation bacteria strains according to claim 1 is characterized in that: the fungi activation medium is the Ma Dingshi substratum in the step 4), and it consists of: glucose 10.0g, peptone 5.0g, KH
2PO
40.5g, MgSO
47H
2O 0.5g; It is dissolved in the 1000mL zero(ppm) water 115 ℃ of moist heat sterilization 15-20min.
10. the screening method of a kind of duomycin degradation bacteria strains according to claim 1 is characterized in that replacing step 1) and step 2 with following steps): 1) bacterial classification enrichment
Gather bacterium slag and waste water, take by weighing waste residue 10g, place the 250mL triangular flask, add 100ml zero(ppm) water, on shaking table, shake up, extract leach liquor; Draw 10mL waste water appearance and bacterium slag leach liquor, insert in the triangular flask of the 250mL that the 100ml enrichment medium is housed respectively, 28 ℃-30 ℃, under the 170r/min concussion condition, secretly cultivated 2-3 days; Get the 5ml nutrient solution then and change in the new identical substratum, under identical culture condition, continue enrichment culture once;
The inorganic salt liquid enrichment medium consists of: glucose 2g, (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA0.015g, agar 13g; Above-mentioned substance is dissolved in the 1000ml zero(ppm) water; Divide and install in the 250mL triangular flask; 115 ℃ of moist heat sterilization 15-20min; In Bechtop, add duomycin, add concentration and be followed successively by 0.1g/L, 1.0g/L, 2.0g/L, 3.0g/L, 4.0g/L, 5.0g/L, 10.0g/L and 15.0g/L, shake up;
2) strains separation
Draw the above-mentioned enrichment culture liquid of 200 μ l, corresponding one by one respectively coating on the inorganic salt solid medium flat board of top said 8 kinds of different chlortetracycline concentrations places under 28 ℃ of-30 ℃ of conditions shading to cultivate 2-7 days;
The inorganic salt solid medium consists of: glucose 2.0g, (NH
4)
2SO
42.0g, K
2HPO
40.5g, KH
2PO
40.5g, MgSO
47H
2O 0.5g, NaCl 0.2g, CaCl
20.1g, FeSO
40.01g, EDTA0.015g, agar 13.0g is dissolved in the 1000ml zero(ppm) water; 115 ℃ of moist heat sterilization 15-20min; Falling before the flat board, in substratum, add duomycin, it is identical with the inorganic salt liquid enrichment medium to add concentration.
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| CN103111037A (en) * | 2013-03-08 | 2013-05-22 | 中国农业科学院农业环境与可持续发展研究所 | Harmless treatment method of chlorotetracycline solid waste |
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| CN103289904A (en) * | 2013-05-14 | 2013-09-11 | 北京理工大学 | Penicillium citrinum LJ318 for degrading chlortetracycline |
| CN103421697A (en) * | 2013-09-02 | 2013-12-04 | 北京理工大学 | Aspergillus oryzae LJ366 strain used for degrading aureomycin |
| CN106434467A (en) * | 2016-10-13 | 2017-02-22 | 南京大学 | High-efficiency antibiotic wastewater treatment agent and preparation method and application thereof |
| CN111495946A (en) * | 2020-04-10 | 2020-08-07 | 苏州市宏宇环境科技股份有限公司 | Restoration method for degrading resistance gene in polluted soil |
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| CN103266064B (en) * | 2013-01-30 | 2015-01-07 | 北京理工大学 | Penicillium oxalicum LJ302 capable of degrading chlortetracycline |
| CN103111037A (en) * | 2013-03-08 | 2013-05-22 | 中国农业科学院农业环境与可持续发展研究所 | Harmless treatment method of chlorotetracycline solid waste |
| CN103111037B (en) * | 2013-03-08 | 2015-08-19 | 中国农业科学院农业环境与可持续发展研究所 | A kind of method for innocent treatment of chlorotetracycline solid waste |
| CN103289904A (en) * | 2013-05-14 | 2013-09-11 | 北京理工大学 | Penicillium citrinum LJ318 for degrading chlortetracycline |
| CN103421697A (en) * | 2013-09-02 | 2013-12-04 | 北京理工大学 | Aspergillus oryzae LJ366 strain used for degrading aureomycin |
| CN106434467A (en) * | 2016-10-13 | 2017-02-22 | 南京大学 | High-efficiency antibiotic wastewater treatment agent and preparation method and application thereof |
| CN106434467B (en) * | 2016-10-13 | 2019-08-06 | 南京大学 | A kind of penicillin wastewater efficient process microbial inoculum and its preparation method and application |
| CN111495946A (en) * | 2020-04-10 | 2020-08-07 | 苏州市宏宇环境科技股份有限公司 | Restoration method for degrading resistance gene in polluted soil |
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