Clostridium and its culturing method and application

A culture method and culture medium technology, applied in the field of microorganisms, can solve the problems of lack of esterase activity and unavailability, and achieve the effect of high culture efficiency

Inactive Publication Date: 2010-04-21
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It can be considered that the rumen is a perfect, natural straw fiber degradation digestion processing plant and efficient anaerobic fermentation tank, and there is no artificial system comparable to it
[0004] Clostridium that can degrade cellulose reported at present has: Clostridium cellulolyticum, isolated from soil, has the ability of decomposing cellulose, but does not have esterase activity (Petitdemange, E., F.Caillet, J.Giallo, andC. Gaudin.1984.Clostridium cellulolyticum sp.nov., a cellular, mesophilic species from decayed grass.Int.J.Syst.Bacteriol.34:155-159); Clostridium cellulolyticum, isolated from soil, grown at high temperature (58°C) , has the ability to decompose cellulose, but cannot utilize xylan and xylose in it (Johnson EA, Sakajoh M, Halliwell G, Madia A, Demain AL.1982.Saccharification of Complex Cellulosic Substrates by the Cellulase System from Clostridium thermocellum.Appl Environ Microbiol. 43(5): 1125-1132)

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Embodiment 1, the separation and preparation of bacterial strain H1

[0031] Take yak rumen content, inoculate it in RC liquid medium with filter paper cellulose as the substrate and subculture to obtain the enrichment; use the enrichment as the inoculum source, inoculate it in the RC liquid medium with cellobiose as the substrate The RC solid medium was separated by the Hungate rolling tube technique.

[0032] It is deposited in the General Microorganism Center (CGMCC) of China Committee for Culture Collection of Microorganisms, and its deposit number is CGMCC No.2125.

Embodiment 2

[0033] Embodiment 2, bacterial strain identification

[0034] 1. Morphological and physiological and biochemical characteristics

[0035] Inoculate the strain H1 (CGMCC No.2125) on RC solid medium (containing 1.5% agar by mass percentage) as the substrate of cellobiose, and roll the tube under the conditions of pH 7.0 and temperature 39°C. Cultured for 2 days.

[0036] Observe the morphological characteristics of colonies and cells, and conduct the following physiological and biochemical experiments: 1. Gram staining test, 2. Contact enzyme test, 3. Indole production test, 4. Urease test, 5. Hydrolysis escin test, 6. Liquefied gelatin test, 7. produce H 2 S test, 8 reduction nitrate test, 9 VP test, 10 substrate utilization test, 11 nitrogen source utilization test.

[0037] The observed and experimental results are as follows:

[0038] 1) Colony characteristics: the diameter of the colony is 1.0-2.0 mm, and the colony is round, with a smooth surface, neat edges, raised, yel...

Embodiment 3

[0050] Embodiment 3, the cultivation of living in the rumen Clostridium H1

[0051] 1) Inoculate Clostridium rumenii H1 at a ratio of 5% in 50 ml RC liquid medium, and culture it at 25° C. and pH 6.

[0052] The composition ( / liter) of RC liquid medium is as follows:

[0053] Rumen fluid: 200ml; Inorganic salt solution I: 100ml; Inorganic salt solution II: 100ml; Cellulose: 1.0g; Cysteine ​​hydrochloride: 0.3g; NaHCO 3 : 5g; resazurin: 0.2mg; distilled water to 10000ml.

[0054] Inorganic salt solution I: K with a mass percent concentration of 0.2% 2 HPO 4 solution.

[0055] Inorganic salt solution II: containing 0.2% KH 2 PO 4 , 0.4% (NH 4 ) 2 SO 4 , 0.4% NaCl, 0.04% MgSO 4 , 0.04% CaCl 2 of aqueous solution.

[0056] Said percentages are all mass percentages.

[0057] The anaerobic environment required for cultivation is achieved by injecting CO 2 The gas is obtained until it reaches 1 atmosphere (1.01×10 5 Pa).

[0058] The experiment was repeated three time...

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Abstract

The present invention discloses one kind of clostridium and its culture process and application. The clostridium is rumen clostridium H1 in the preservation number of CGMCCNo.2125, and has high culture efficiency and contained cellulose degrading composite enzyme system comprising exocellulase, endo cellulase, xylanase, mannanase, esterase and pectase. The rumen clostridium H1 has capacity of decomposing natural cellulose, esterase activity of decomposing ester, easy medium temperature culture at 39 deg.c and capacity of utilizing pentose. It may find its wide application in cellulose degrading industry.

Description

technical field [0001] The invention relates to a clostridium in the field of microorganisms, a culture method and application thereof. Background technique [0002] Plants produce organic matter through photosynthesis, 35% or more of which exist in the form of cellulose substances such as crop straws, but such cellulose substances have not been fully developed and utilized, and thus become an important source of environmental pollution. Cellulosic substances can be degraded into five-carbon sugars and six-carbon sugars, and then the sugars can be fermented by microorganisms to produce fuel alcohol or other raw materials for chemical products. Therefore, people have been exploring and researching the conversion of cellulosic waste into renewable energy. At present, chemical or physical methods are generally used to hydrolyze cellulose to produce sugar, and then bio-fermented sugar to produce fuel ethanol, but this technology has high costs and has not been industrialized on...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C12P1/04
Inventor 东秀珠蔡世淳张科贵
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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