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Microbial strain expressing glycosidase and application technique thereof to biological enzyme papermaking

A thermophilic glucosidase and papermaking pulp technology, applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve problems such as poor thermal stability and unstable organic solvents

Inactive Publication Date: 2013-07-24
SHENZHEN POLYTECHNIC +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, most of the glycosidases that have been reported belong to normal temperature enzymes. Due to their poor thermal stability and instability to organic solvents, they are not suitable for enzymatic synthesis of glycoside compounds. Therefore, it is necessary to look for new and stable , high temperature resistant enzyme sources have gradually become the focus of attention

Method used

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  • Microbial strain expressing glycosidase and application technique thereof to biological enzyme papermaking
  • Microbial strain expressing glycosidase and application technique thereof to biological enzyme papermaking
  • Microbial strain expressing glycosidase and application technique thereof to biological enzyme papermaking

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] [Example 1] Using crops and leaves as the ratio screening of papermaking raw material pulp

[0042] In this embodiment, the alkaloid sodium ethoxide is used, and the raw materials are corn stalks, wheat stalks, and banana leaves, and a ratio experiment of different raw materials is carried out. The purpose of raw material slurry ratio screening is to explore for the subsequent enzymatic papermaking and reduce papermaking pollution.

[0043] The specific proportioning schemes are shown in Table 1 to Table 4 below. The ratio of corn stalks: wheat is from 1:1 to 10:1; banana leaves: fishbone leaves are from 4:1 to 9:1; 1:1 to 5:1.

[0044] Table 1: Biological enzymatic papermaking experiment with the ratio of two crops

[0045]

[0046] Table 3 Proportion of crops and leaves for bio-enzymatic papermaking experiment (corn stalks and banana leaves)

[0047]

[0048] The results of the bio-enzymatic papermaking experiment with the ratio of the above two crops show t...

Embodiment 2

[0049] [Example 2] Screening of technological conditions of enzymatic method combined with organic alkali papermaking method

[0050] The cellulase used in this example: the model is 1.4 million U / g, the batch number is 20120280008YX, and the manufacturer is Xiasheng Industrial Group Co., Ltd.

[0051] β-glucanase: the model is 30 million U / g, the batch number is 20120280008YX, and the manufacturer is Xiasheng Industrial Group Co., Ltd.

[0052] The results of the research on the environmental protection process conditions of enzymatic method + organic alkali are shown in Table 5:

[0053] Table 5: Exploration of enzymatic papermaking conditions

[0054]

[0055] It can be seen from Table 5 that the required action time is also significantly different when the concentration of the enzyme is different. According to the cost, the group with a smaller amount of enzyme and a shorter action time is preferred. Because the amount of enzyme added is directly related to the produc...

Embodiment 3

[0056] [Example 3] Construction of a vector expressing a biological enzyme gene

[0057] The gene thermophilic glycosidase 0602, also called TN0602 in this paper, was extracted from the genome of Thermotoga naphthophila (Thermotoga naphthophila, Japan Collection of Microorganisms, RIKEN BioResource Center) by conventional PCR. Ferment hyperthermophilic archaea, collect the bacteria by centrifugation, extract the gene with a kit, transfer the thermophilic glycosidase 0602 gene from the genome by PCR, and then transfer it into Escherichia coli (E.coli BL21 (DE3)) for further Expression, purification and characterization of basic enzymatic properties.

[0058] Experimental materials and methods

[0059] Genome extraction: Escherichia coli cells transformed with the thermophilic glycosidase 0602 gene were collected by centrifugation, and their genomes were extracted with a kit (Hangzhou Viteje Co., Ltd.) according to the instructions of the kit.

[0060] Primer design: Using the...

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Abstract

The invention provides a method for decomposing papermaking pulp by using engineering bacteria secreting thermophilic glycosidase. The technique comprises the following steps: 1) using crop straws as a raw material, crushing and soaking for 20-30 h to form a raw slurry; 2) amplifying engineering bacterium seed secreting thermophilic glycosidase, conducting a first stage culture and a second stage culture, and inducing expression of the thermophilic glycosidase at 25 DEG C-37 DEG C with IPTG of 0.2 0.7mM when OD value of a second strain reaches 0.4-0.8; 3) adding a second stage strain liquid obtained in the step 2) according to a volume ratio of 1:20-40 into raw slurry in the step 1), and treating the slurry at room temperature overnight; and 4) fishing out the slurry, boiling, washing, crushing, conducting papermaking and baking for molding. The microbial strain secreting glycosidase provided by the invention has decomposition effect on papermaking raw pulp similar to that of a commonly used enzyme reagent, and the cost is greatly reduced.

Description

technical field [0001] The invention relates to the technical field of papermaking by biological enzyme method, in particular to a microbial strain expressing glycosidase and its technology for papermaking. Background technique [0002] Due to the rapid development of biotechnology, the advantages of other industries benefiting from its development results are becoming more and more obvious and spreading rapidly. The paper industry is well known as a major polluting industry, especially in the process of pulping and papermaking that produces a large amount of chemical pollution. According to the statistical data in 2008, China's pulp and paper production reached 79.8 million tons, becoming the largest paper country in the world, and the country also spent a lot of money on the treatment of sewage, waste gas and solid waste in the paper industry . Applying modern biotechnology to energy saving, emission reduction and pollution control in the paper industry is the most promi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): D21C5/00C12N1/21C12R1/19
Inventor 王妍吴泽旋陈莫喇文军李云高仁钧解桂秋孔繁思程金
Owner SHENZHEN POLYTECHNIC
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