Acidic trehalase TreA and gene and application thereof

A trehalase and gene technology, which is applied in the field of agricultural genes, can solve the problems such as the inability of seaweed enzymes to meet the needs of modern industries, and achieve the effects of high specific activity and excellent performance.

Inactive Publication Date: 2018-11-20
INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to solve the problem that the existing seaweed enzymes cannot meet the needs of modern industries, the present invention provides an acid trehalase derived from fungi, which has excellent enzymatic properties and can meet the requirements of modern industries

Method used

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  • Acidic trehalase TreA and gene and application thereof
  • Acidic trehalase TreA and gene and application thereof
  • Acidic trehalase TreA and gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Example 1 Obtaining Trehalase TreA

[0053] 1. Cloning trehalase coding gene TreA

[0054]Centrifuge the Bispora sp.MEY-1 bacteria cultured in liquid for 3 days at 12,000rpm for 10min, put the collected mycelium into a high-temperature sterilized mortar, grind it quickly with liquid nitrogen to powder, and then transfer the ground bacteria Transfer to a new 50mL centrifuge tube containing 15ml CTAB lysate, gently invert up and down to mix well, place in a 65°C water bath for 3 hours, and invert up and down gently once every 20min to fully lyse the bacteria. Centrifuge at 12,000 rpm at 4°C for 10 min, pipette the supernatant into a new centrifuge tube, add an equal volume of chloroform for extraction, and place at room temperature for 5 min. Centrifuge at 12,000 rpm for 10 min at 4°C. Take the supernatant and add an equal volume of phenol / chloroform for extraction, and place it at room temperature for 5 minutes. Centrifuge at 12,000 rpm for 10 min at 4°C. In order to...

Embodiment 2

[0073] Example 2 Determination of Partial Properties of Trehalase TreA

[0074] The activity analysis of the trehalase of the present invention is carried out by DNS method. The specific method is as follows: at pH 4.0, 50°C, 1 mL of reaction system includes 100 μL of appropriate diluted enzyme solution, 900 μL of substrate, react for 10 min, add 1.5 mL of DNS to terminate the reaction, and boil for 5 min. After cooling, the OD value was measured at 540 nm. Definition of trehalase activity unit: Under certain conditions, the amount of enzyme needed to decompose trehalose to generate 1 μmol reducing sugar per minute is 1 activity unit (U).

[0075] 1. The optimal pH and pH stability of Trehalase TreA

[0076] At different pH, the enzymatic reaction was carried out to determine the optimum pH of trehalase TreA. The buffers used are pH 1.5-4.0 KCl-hydrochloric acid buffer, pH 4.0-7.0 citric acid-disodium hydrogen phosphate buffer series. Purified trehalase TreA in different p...

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Abstract

The invention belongs to the technical field of agricultural genes and in particular relates to acidic trehalase TreA sourced from a fungus and a gene and application thereof. The amino acid sequenceof the trehalase TreA is as shown in SEQ ID NO.1 or SEQ ID NO.2. The invention provides a novel trehalase gene. The trehalase encoded by the trehalase gene has high properties and can be applied to industries of insect control, foot, starch sugar, medicines and the like.

Description

technical field [0001] The invention belongs to the technical field of agricultural genes, and in particular relates to an acid trehalase derived from fungi and its gene and application. Background technique [0002] Trehalose is a non-reducing disaccharide formed by connecting two molecules of glucose with α,α-1,1-glycosidic bonds, and its molecular formula is C 12 h 22 o 11 , with a molecular weight of 342.31, usually in the form of dihydrates. Trehalose exists widely in nature, including plants, algae, fungi, bacteria, insects and other species, and is decomposed into glucose by trehalase to provide energy for the body. [0003] The sources of trehalase are mainly divided into three categories: the first category comes from bacteria; the second category comes from plants and animals; the third category comes from fungi. According to the optimum pH value of the enzyme-catalyzed reaction, trehalase derived from fungi can be divided into two types: one is acid trehalase ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/24C12N15/56C12N15/81C12N1/19C12P19/14C12P19/02C12R1/84
CPCC12N9/2402C12N15/815C12P19/02C12P19/14C12Y302/01028
Inventor 罗会颖姚斌蒋肖涂涛黄火清苏小运王亚茹王苑柏映国孟昆
Owner INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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