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Method for catalytically synthesizing starch xanthate with yeast display lipase

A technology of xanthate and synthetic starch, applied in the field of bioengineering, can solve the problems of limited commercial application, high production cost, complicated and time-consuming immobilization process, etc., to achieve improved operational stability, low production cost, and shortened reaction time Effect

Inactive Publication Date: 2014-07-16
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Lipase is currently the most widely used enzyme in esterification synthesis, but its commercial application is greatly limited by the high production cost and complicated and time-consuming immobilization process.

Method used

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  • Method for catalytically synthesizing starch xanthate with yeast display lipase
  • Method for catalytically synthesizing starch xanthate with yeast display lipase

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1 Preparation of Yeast Displaying Lipase

[0019] Synthesize the lipase gene (Genbank number: AF229435) of Rhizopus oryzae and the cell wall α-lectin gene of Pichia pastoris GS115 (Genbank number: M28164) by artificial synthesis, and add Connect the peptide sequence GSSGGSGGSGGSGGSGS(linker), and get the nucleotide sequence pro-ROL-linker-α-agglutinin after connection, and add EcoR I and Not I restriction sites at both ends of the sequence, where pro-ROL is the lipase gene , α-agglutinin is the cell wall α lectin gene.

[0020] Using the above artificially synthesized sequence as a template, PCR amplification was performed using the following primer pair,

[0021] Upstream primer: 5'-AAGGAAAAAAAGAATTCGTTCCAGTTTCTGG-3';

[0022] Downstream primer: 5'-TTTTCCTTTTGCGGCCGCTAATGAAACG-3'

[0023] The PCR reaction system is: 1 μl of template DNA, 0.5 μl of high-fidelity DNA polymerase, 0.4 μl of dNTP (50 mM), 0.5 μl of upstream and downstream primers, 5 μl of 10×PCR ...

Embodiment 2

[0027] Example 2 Yeast shows lipase catalyzed synthesis of starch xanthate

example 1

[0028] Example 1 Take 350g of corn starch and prepare 35% starch milk (starch dry basis) with water, absorb water at 65°C for 15 minutes to make the starch milk swell with water, add 1g of the above-mentioned prepared yeast to display lipase after cooling, and then add 10g of CS in batches 2 , placed in a 85-1 type magnetic stirrer and stirred to start the reaction, the rotation speed was 200 rpm, the reaction temperature was kept at 46°C, the stirring was stopped after 1 hour of reaction, the precipitate was collected by centrifugation, and the precipitate was washed with water for many times to wash away unreacted Acetic anhydride, and then the precipitate is dried and crushed to obtain the finished product.

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Abstract

The invention discloses a method for catalytically synthesizing starch xanthate with yeast display lipase. The method comprises the following steps: dissolving starch of grain in water; adding CS2 and yeast display lipase after imbibition; stirring the mixture to react at 45-55 DEG C for 1-1.5 hours; and carrying out separation and purification to prepare the starch xanthate, wherein the yeast display lipase is prepared by transforming recombinant plasmid which is subjected to linearization into Pichia pastoris GS115, inoculating the obtained transformant in a BMMY culture medium, carrying out inducing culture for 72-144 hours, centrifuging to collect a thallus, and carrying out washing, bio-imprinting and freeze-drying on the thallus. By displaying the lipase outside the cell and utilizing the lipase preparation to catalytically synthesize the starch xanthate, the invention improves the transformation efficiency, shortens the reaction time and lowers the production cost.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to a method for synthesizing starch xanthate catalyzed by yeast display lipase. Background technique [0002] Starch xanthate is starch and CS 2 A starch derivative obtained by xanthation reaction under alkaline conditions. It can adsorb various heavy metal ions in wastewater under acidic conditions, and is often used in the recovery of heavy metal ions from electroplating wastewater. As a traditional method for purifying wastewater, starch xanthate not only has low energy consumption, high efficacy, rapid response, and easy operation. Moreover, heavy metals can be effectively recovered, and economic and social benefits can be obtained while obtaining environmental benefits. [0003] Compared with the chemical synthesis of starch xanthate, the biological enzymatic method can obtain a reaction product with a high degree of substitution in a short period of time. For the sam...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P19/04C12N15/63C12N1/19C12N9/20C12R1/84
Inventor 阮晖王睿之古再丽努尔徐娟周陈伟林吉恒何国庆
Owner ZHEJIANG UNIV
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