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Phosphorus-dissolving pseudomonas putida L13 and fermentation process thereof

A technology of Pseudomonas putida and phosphorus solubilization, applied in the field of microorganisms, can solve the problems of complex fermentation production process and narrow practical application range, and achieve strong application prospects, strong environmental adaptability, and growth-promoting effects

Inactive Publication Date: 2012-08-29
TOBACCO RES INST OF HUBEI PROVINCE +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] Before this application was filed, Huazhong Agricultural University disclosed Clostridium butyricum A5-4 and its application on October 6, 2010 (patent publication number: CN101851596A, patent application number: 2010101534192, patent application date: 2010 April 21, 2010), the Clostridium phosphobutyricum A5-4 is an anaerobic bacterium, which is mainly used in sewage treatment in an anaerobic environment and as a growth-promoting bacterium in the flooded state of rice. Since this bacterium can only be used In an anaerobic environment, its practical application range is narrow, and the fermentation production process is relatively complicated

Method used

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  • Phosphorus-dissolving pseudomonas putida L13 and fermentation process thereof
  • Phosphorus-dissolving pseudomonas putida L13 and fermentation process thereof
  • Phosphorus-dissolving pseudomonas putida L13 and fermentation process thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Embodiment 1 Shake flask fermentation culture

[0059] A. First-class seed preparation: take the slanted seeds of Pseudomonas phospholytica putida L13 (preservation number is CCTCC NO: 2010342) and draw a line on the LB medium plate, put it into a 28° C. incubator for 14 hours, and free single colonies appear. This is the primary seed;

[0060] B. Preparation of secondary seeds: use an inoculation loop to take a single colony of the primary seed of step A, and insert it into a 250mL shake flask with 50mL of LB liquid medium, control the temperature at 28°C, and the shaker speed at 180rpm, and ferment and cultivate 12h, OD 600 Reaching 2.0, this is the second-level seed;

[0061] C, fermentation culture: get the secondary seed liquid of step B to insert in the 250mL shaking flask that 30mL fermentation medium is housed with the inoculum size of 2% (V / V) and carry out fermentation culture, control shaker temperature to be 28 ℃, rotating speed 180rpm, fermentation cultu...

Embodiment 2

[0066] Embodiment 2 Shake flask fermentation culture

[0067]A. First-class seed preparation: take the slanted seeds of Pseudomonas phospholytica putida L13 (preservation number is CCTCC NO: 2010342) and draw a line on the LB medium plate, put it into a 28° C. incubator for 14 hours, and free single colonies appear. This is the primary seed;

[0068] B. Preparation of secondary seeds: use an inoculation loop to take a single colony of the primary seed of step A, and insert it into a 250mL shake flask with 50mL LB liquid medium, control the temperature at 30°C, and shake the table at a speed of 200rpm, and ferment and cultivate 14h, OD 600 Reaching 2.0, this is the second-level seed;

[0069] C, fermentation culture: get the secondary seed liquid of step B to insert in the 250mL shaking flask that 40mL fermentation medium is housed with the inoculum size of 3% (V / V) and carry out fermentation culture, control shaker temperature to be 30 ℃, rotating speed 200rpm, fermentation...

Embodiment 3

[0074] Embodiment 3 Shake flask fermentation culture

[0075] A, primary seed preparation: take the slant seeds Pseudomonas putida L13 (preservation number is CCTCC NO: 2010342) and draw a line on the LB medium plate, put it into a 28°C incubator and cultivate it for 10h, free single colonies appear, This is the primary seed;

[0076] B. Preparation of secondary seeds: use an inoculation loop to take a single colony of the primary seed of step A, and insert it into a 250mL shake flask with 30mL LB liquid medium, control the temperature at 26°C, and shake the table at a speed of 160rpm. Fermentation 10h, OD 600 Reaching 2.0, this is the second-level seed;

[0077] C, get the secondary seed liquid of step B to insert in the 250mL shaking flask that 20mL fermentation medium is housed with the inoculum size of 0.5% (V / V) and carry out fermentation culture, control shaker temperature is 26 ℃, and rotating speed is 160rpm, Fermentation culture to 36h, OD 600 reached 15.0;

[00...

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Abstract

The invention belongs to the technical field of agricultural microbiology, and particularly relates to the separation screening and fermentation cultivation of phosphorus-dissolving pseudomonas putida L13. A phosphorus-dissolving pseudomonas putida L13 strain with the capacity of dissolving phosphorus is obtained through separation, and the preservation No. is CCTCC No.2010342. The fermentation process comprises the following steps of: (1) taking L13 bacterial strain slope seeds for streak cultivation on an LB (lysogeny broth) culture medium flat plate to obtain first-level seeds; (2) inoculating a first-level seed single bacterial colony into an LB culture medium shake flask to make OD (optical density) 600 be 2.0 to obtain second-level seeds; (3) inoculating the second-level seeds into a shake flask for fermentation cultivation to obtain the pseudomonas putida; or inoculating the second-level seeds into a fermentation tank so as to obtain the pseudomonas putida through optimized cultivation. The pseudomonas putida strain disclosed by the invention grows rapidly, the yield of the pseudomonas putida is high, and the viable count can reach more than 10 billions. The phosphorus-dissolving amount of the phosphorus-dissolving pseudomonas putida in a PKO (Pikovskaya') culture medium in which 1-3g / L of calcium phytate is the unique phosphorus source can reach 120-220mg / L, and the phosphorus-dissolving amount of the phosphorus-dissolving pseudomonas putida in a PKO culture medium in which 4-6g / L of tricalcium phosphate is the unique phosphorus source can reach 300-500mg / L.

Description

technical field [0001] The invention relates to the field of microorganisms, in particular to a strain of Pseudomonas putida phospholyticum and its fermentation process. Background technique [0002] Only 1% to 5% of the total phosphorus source in the soil exists in the form of soluble phosphorus that can be used by plants. Due to the adsorption effect of the soil, about 90% of traditional chemical fertilizers will be fixed by the soil soon after being applied to the soil and become insoluble phosphorus that cannot be used by plants, and only a small part of the soluble phosphorus that can finally exist in a stable form. Therefore, after the 1980s, with the continuous development of agriculture, the soluble phosphorus in the soil decreased to a considerable extent, while the reduction in the total inorganic phosphorus sources in the soil was relatively small. [0003] There are a variety of beneficial microbial flora in the rhizosphere of crops, we call it plant growth prom...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C05F11/08C12R1/40
Inventor 陈守文王昌军信健冀志霞李进平祖秉桥祁高富马昕杨树李建平
Owner TOBACCO RES INST OF HUBEI PROVINCE
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