Mannanases and recombinant expression bacterial strain thereof

A technology of mannanase and engineering bacteria, which is applied in the field of microbial engineering and can solve problems such as intolerance to pepsin

Active Publication Date: 2013-12-18
QINGDAO VLAND BIOTECH GRP
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Problems solved by technology

Although the mannanase produced by Trichoderma and Penic

Method used

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  • Mannanases and recombinant expression bacterial strain thereof
  • Mannanases and recombinant expression bacterial strain thereof
  • Mannanases and recombinant expression bacterial strain thereof

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[0026] The method of the present invention will be further explained below in conjunction with examples. The experimental methods without specific conditions in the examples can usually be under conventional conditions, as described in the "Molecular Cloning Experiment Guide" compiled by J. Sambrook (Sambrook) et al. Or run under the conditions recommended by the manufacturer. Those skilled in the art can better understand and master the present invention with the help of the embodiments. However, the protection and scope of the claims of the present invention are not limited to the cases provided.

[0027] 1 Cloning of the mannanase gene from Penicillium decumbens

[0028] 1.1 Extraction of total DNA

[0029] Culture Penicillium decumberns (P. decumberns) overnight, place an appropriate amount of bacteria in a centrifuge tube, centrifuge at 13000 rpm for 5 min, discard the supernatant; add 400μl extraction buffer (100 mM Tris-HCl, 100 mM EDTA, 250 mM NaCl, 1% SDS); then add 100mg...

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Abstract

The invention relates to mannanases and a recombinant expression bacterial strain thereof. Expression genes of the mannanases is cloned from penicillium decumbens. Amino acid sequences of the mannanases are shown in the formula of SEQ ID NO: 1. The invention also comprises Pichia pastoris pdman-2 engineering bacteria and Trichoderma reesei pdman-2 engineering bacteria for expression of the mannanases. The novel expression genes of the mannanases are cloned from penicillium decumbens, and the Pichia pastoris pdman-2 engineering bacteria and the Trichoderma reesei pdman-2 engineering bacteria for expression of the novel expression gene are constructed. The mannanase expressed by the Pichia pastoris pdman-2 engineering bacteria has the most suitable pH value of 4.5, the most suitable temperature of 55 DEG C and stable enzyme activity in a pH value of 2.0-7.0, and can resist gastric acid and pepsin. The mannanase expressed by the Trichoderma reesei pdman-2 engineering bacteria has the most suitable pH value of 4.5, the most suitable temperature of 55 DEG C and stable enzyme activity in a pH value of 3.5-8.0, and can resist trypsin. The mannanases have feeding-enzyme application values.

Description

technical field [0001] The invention belongs to the technical field of microbial engineering, and in particular relates to a mannanase and its recombinant expression strain. Background technique [0002] In nature, plant cell walls are mainly composed of cellulose, hemicellulose, and lignin. Mannan is an important component of plant hemicellulose, the second largest renewable hemicellulose carbohydrate after cellulose, which is connected by β-1,4-D-mannose to form a linear polysaccharide. There are mainly substituent groups such as glucosyl, acetyl and galactosyl on the side chain of the polysaccharide. Mannan is hydrophilic, absorbs a large amount of water in the digestive tract of monogastric animals, increases the viscosity of the contents of the digestive tract, thereby resisting gastrointestinal peristalsis, and directly affects the digestion and absorption of nutrients by animals. In recent years, with the widespread application of soy products (soybean meal, etc.) i...

Claims

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Application Information

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IPC IPC(8): C12N9/42C12N15/56C12N15/63C12N1/15C12N1/19A23K1/165C12R1/84C12R1/885
Inventor 黄亦钧程斯达王华明刘鲁民曲音波刘国栋陈梅
Owner QINGDAO VLAND BIOTECH GRP
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