Method for preparing protoplast of white rot fungi

A technology of protoplasts and white-rot fungi, applied in biochemical equipment and methods, methods based on microorganisms, fungi, etc., can solve the problems of difficult formation of dominant populations and slow colonization, and achieve easy operation and simple preparation process Effect

Inactive Publication Date: 2010-05-26
TIANJIN INST OF ANIMAL HUSBANDRY & VETERINARY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the natural state, due to the slow colonization of white rot fungi, it is difficult to form a dominant population, which requires stricter fermentation conditions. In order to breed mutant strains with higher lignin degradation ability and strengthen lignin degradation, it is necessary to Its protoplast preparation method was studied

Method used

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Examples

Experimental program
Comparison scheme
Effect test

example 1

[0014] Connect the robust mycelium on the PDA solid medium and cultivate it in the liquid medium for 240h, pretreat the mycelia with 0.1% β-mercaptoethanol, and use mixed enzymes (lysozyme: cellulase: the optimal concentration ratio of helicase is 2:2:1; enzyme solution pH4) Enzymolysis at 28°C for 3h; use 0.6mol / L MgSO 4 When used as an osmotic pressure stabilizer, the number of protoplasts reached 6×10 6 individual / mL.

example 2

[0016] Connect the robust mycelium on the PDA solid medium and cultivate it in the liquid medium for 288h, pretreat the mycelium with 0.3% β-mercaptoethanol, and adopt mixed enzymes (lysozyme: cellulase: the optimum concentration ratio of helicase is 2:2:1; enzyme solution pH5,); enzymolysis at 32°C for 3h; use 0.6mol / L MgSO 4 When used as an osmotic pressure stabilizer, the number of protoplasts reached 7.2×10 6 individual / mL.

example 3

[0018] Connect the robust mycelium on the PDA solid medium and cultivate it in the liquid medium for 264h, pretreat the mycelia with 0.1% β-mercaptoethanol, and use mixed enzymes (lysozyme: cellulase: the optimum concentration ratio of helicase is 2:2:1; enzyme solution pH6); enzymatic hydrolysis at 30°C for 3h; use 0.6mol / L MgSO 4 When used as an osmotic pressure stabilizer, the number of protoplasts reached 8×10 6 individual / mL.

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Abstract

The invention provides a method for preparing protoplast of white rot fungi. The method comprises the following steps: inoculating strong mycelium on a PDA solid medium to a liquid medium and culturing for 240 to 290 hours; pretreating the mycelium by 0.1 to 0.3 percent beta-mercaptoethanol; performing enzymolysis on the mycelium at the temperature of between 26 and 32 DEG C for 1 to 4 hours by adopting mixed enzyme of muramidase, cellulase and helicase with the pH of between 4 and 7; and ensuring that the number of the protoplast reaches 5-8*10<6> per milliliter when 0.6mol / L MgSO4 is used as an osmotic pressure stabilizer. The method for preparing the protoplast of the white rot fungi has the advantages of simple preparation process and easy operation. The preparation yield of the protoplast reaches 5-8*10<6> per milliliter.

Description

technical field [0001] The invention belongs to a method for breeding microorganisms in bioengineering, in particular to a method for preparing protoplasts of white-rot fungi. Background technique [0002] Lignin is the main component of plant cell walls. There is almost no enzyme to degrade lignin in the digestive tract of livestock and poultry. Herbivores can partially degrade lignin by parasitic microorganisms in the digestive tract, but monogastric animals do not have this ability. Lignin is the biggest obstacle for animals to digest plant-based feed, and microbial fermentation is the most ideal way to overcome this problem. Breeding microbial strains with high lignin degradation rate and easy fermentation on feed is a key technology for the aquaculture industry to realize "grain saving". White-rot fungi are the most effective microorganisms for degrading lignin, and they are also the model strains often used in lignin degradation research. However, in the natural stat...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/14C12R1/645
Inventor 范寰廖伟
Owner TIANJIN INST OF ANIMAL HUSBANDRY & VETERINARY
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