In-situ enriching method and screening method of pullulanase indigenous strains

An in situ enrichment, pullulanase technology, applied in biochemical equipment and methods, determination/inspection of bacteria, microorganisms, etc. Easy to operate, clear purpose, reliable effect

Active Publication Date: 2018-01-05
HENAN UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The traditional enrichment method of pullulanase-producing indigenous strains can generally isolate a small number of strains, but the existing problems are also obvious. These deficiencies are concentrated in: (1) The isolation method is too programmed and single, and the isolated strains The repetition rate is extremely high, the diversity is poor, and there are few innovative research results; (2) Enrichment culture for a certain period of time before isolation culture is obviously more targeted than no enrichment, and the effect is slightly better, but in vitro enrichment also has The enriched environment of the sample still has natural defects that are fundame

Method used

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  • In-situ enriching method and screening method of pullulanase indigenous strains
  • In-situ enriching method and screening method of pullulanase indigenous strains

Examples

Experimental program
Comparison scheme
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Example Embodiment

[0026] Example 1

[0027] The in situ enrichment method of the indigenous pullulanase-producing strain in this embodiment includes:

[0028] A. Substrate handling

[0029] Take 5g of sterilized absorbent cotton in the ultra-clean workbench and put it into a sterilized beaker containing 15g of pullulan and 50mL of sterile water, and turn the absorbent cotton to make the pullulan infiltrate and adhere to the absorbent cotton as much as possible. Absorbent cotton blocks are kept sterile.

[0030] B. In-situ enrichment in soil

[0031] Dig a 5cm deep pit in the sampling environment soil (the wasteland soil on both sides of the Luohe River in the suburbs of Luoyang), sprinkle 100mL sterile water in the pit, take out the absorbent cotton block processed in step A, and arrange the cotton block into a rectangle ( subject to no breakage), lay flat on the bottom of the hole, and cover the absorbent cotton with the in-situ soil when digging the hole. After 8 days of in-situ enrichmen...

Example Embodiment

[0035] Example 2

[0036] The in situ enrichment method of the indigenous pullulanase-producing strain in this embodiment includes:

[0037] A. Substrate handling

[0038] Take 5g of sterilized absorbent cotton in the ultra-clean workbench and put it into a sterilized beaker containing 20g of pullulan and 50mL of sterile water, and turn the absorbent cotton to make the pullulan infiltrate and adhere to the absorbent cotton as much as possible. Absorbent cotton blocks are kept sterile.

[0039] B. In-situ enrichment in soil

[0040]Dig a 10cm deep pit in the sampling environment soil (corn field soil in the suburbs of Luoyang), sprinkle 150mL sterile water in the pit, take out the absorbent cotton block processed in step A, and arrange the cotton block into a rectangle ( subject to no breakage), lay flat on the bottom of the hole, and cover the absorbent cotton with the in-situ soil when digging the hole. After 12 days of in-situ enrichment, the absorbent cotton block was c...

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Abstract

The invention relates to an in-situ enriching method and a screening method of pullulanase indigenous strains. The in-situ enriching method comprises the following steps: burying a matrix to which amylopectin is adhered into soil; enriching; then taking out the matrix; and collecting soil on the surface of the matrix to obtain the strain-enriched soil. The screening method comprises the followingsteps: uniformly mixing the strain-enriched soil and sterile water; centrifuging; performing gradient dilution on an upper suspension; coating a flat separating plate with the gradient diluting solution; culturing; and then identifying. The enriching method has the major characteristic that amylopectin is used as a primary inducing substrate for in-situ enriching the pullulanase indigenous strainsin soil, so that the quantity of pullulanase indigenous populations can be increased in an original ecological environment, and particularly some weak populations with pullulanase activity can be certainly proliferated, and as a result, the detection rate of the target strains can be increased.

Description

technical field [0001] The invention relates to an in-situ enrichment method and a screening method for an indigenous strain producing pullulanase, and belongs to the technical field of bacterial enrichment and screening. Background technique [0002] Microorganisms are an important part of biological resources and material cycles in nature. It is generally believed that the types of microorganisms known to people are only 1%-10% of the actual stock, and some people even think that they are less than 0.1%. It is a long and arduous job. Starch is a polysaccharide compound that is consumed in a huge amount in people's lives. Natural starch is composed of 10%-30% amylose and 70%-90% amylopectin, and the utilization rate of starch mainly depends on the amylopectin The level of utilization. The α-1,6 glycosidic bonds contained in amylopectin are difficult to hydrolyze, which becomes a bottleneck problem in the efficient utilization of starch, and pullulanase exclusively hydroly...

Claims

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Application Information

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IPC IPC(8): C12N1/02C12N1/20C12Q1/04
Inventor 宋月芹孙会忠侯小改宋鹏
Owner HENAN UNIV OF SCI & TECH
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