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A Saccharomyces cerevisiae strain expressing xylose isomerase and its construction method

A technology of Saccharomyces cerevisiae strain and construction method, which is applied in the field of Saccharomyces cerevisiae strains, can solve the problems of not being able to meet the fermentation requirements and the slow growth rate of recombinant bacteria, and achieve the effects of high ethanol yield and fast xylose fermentation

Active Publication Date: 2020-08-04
SICHUAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The purpose of the invention is to solve the existing xylA After the gene is heterologously expressed in Saccharomyces cerevisiae, the growth rate of the recombinant bacteria constructed by it is relatively slow in xylose, which cannot meet the needs of fermentation; a strain based on the xylose isomerase pathway and capable of solving the above problems is provided. Industrial Saccharomyces cerevisiae Efficiently Fermenting Xylose to Ethanol

Method used

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  • A Saccharomyces cerevisiae strain expressing xylose isomerase and its construction method
  • A Saccharomyces cerevisiae strain expressing xylose isomerase and its construction method

Examples

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Embodiment 1

[0026] A strain of Saccharomyces cerevisiae expressing xylose isomerase, the Saccharomyces cerevisiae ( Saccharomyces cerevisiae SEB7) strains are deposited in the China General Microorganism Culture Collection and Management Center, and the preservation number is CGMCC11327. The specific construction method of this Saccharomyces cerevisiae strain SEB7 is as follows:

[0027] 1) Construct the starting strain:

[0028] Saccharomyces cerevisiae SEB3 was cultured on sporulation medium (0.5 g / L glucose, 20 g / L potassium acetate, 2 g / L yeast powder, pH 5.5) for 2 days, and then treated with lysozyme at 30°C for 10-20 min Then, single spores were picked on a tetraspore analyzer, and haploid cells were obtained after verification. The present invention only selects the haploid whose sex is α, and finally selects the haploid strain SEB3α25.

[0029] Using the plasmid pBlu-LTKTL-TDH3 as the template, the primers Fg3 / Rg3 were used to amplify the loxP-KanMX-loxP The fragment was ...

Embodiment 2

[0057] This example is a comparative example of Example 1. The construction steps and construction conditions in this example are the same as those in Example 1. The only difference is that primers PXYLA1-F / PXYLA1-R and PXYLA2-F / are used in step 2.4). PXYLA2-R converts the original gene sequence of xylose isomerase PXYLA1 Amplified, it and plasmid pRS426-P TDH3 -T TDH3 through EcoR After I digestion, ligated with T4 ligase, after digestion and sequencing verification, the multi-copy expression vector pRS426-PXYLA1 was obtained.

[0058] Saccharomyces cerevisiae YCPA1 was obtained after construction through step 3). The strain YCPA1E was obtained by carrying out the aerobic growth acclimation and micro-aerobic fermentation acclimation of the strain YCPA1 in step 4).

[0059] By evaluating the growth of YCPA1 and YCPA2, the strains needed to be cultured for 36 h before reaching the mid-logarithmic growth stage before the fifth passage. In the first 5 generations of domes...

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Abstract

The present invention discloses a strain of Saccharomyces cerevisiae expressing xylose isomerase and its construction method, which solves the problem of existing xylA After the gene is heterologously expressed in Saccharomyces cerevisiae, the growth rate of the recombinant bacteria constructed by it is relatively slow in xylose, which cannot meet the needs of fermentation. The present invention includes said Saccharomyces cerevisiae ( Saccharomyces cerevisiae The SEB7) strain is preserved in the China General Microorganism Culture Collection and Management Center, and the preservation number is CGMCC11327. The invention also provides the construction method of the Saccharomyces cerevisiae SEB7. The bacterial strain SEB7 that the present invention obtains can utilize 16.95g / L xylose to produce the ethanol of 6.98 g / L in the fermentation time of 48 h, and ethanol yield reaches 0.412 g / g xylose; This bacterial strain has xylose fermentation fast, The advantages of high ethanol yield.

Description

technical field [0001] The invention relates to a yeast strain, in particular to a Saccharomyces cerevisiae strain expressing xylose isomerase and a construction method of the strain. Background technique [0002] In recent years, fuel ethanol has received extensive attention as one of the most promising clean energy sources. Lignocellulose, which uses agricultural and forestry wastes as raw materials, is regarded as an ideal raw material for fuel ethanol production due to its wide sources, large reserves, low price, and renewable advantages. Saccharomyces cerevisiae( Saccharomyces cerevisiae ) is the most commonly used microorganism in industrial ethanol production due to its advantages of fast glycolysis, high ethanol yield, and good environmental tolerance. However, in the sugar solution obtained by hydrolysis of lignocellulose, about 20%–30% of the sugar is xylose, which is because wild-type Saccharomyces cerevisiae cannot directly use xylose for growth and fermentatio...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/19C12N15/61C12N15/81C12R1/865
Inventor 汤岳琴李云成苟敏孙照勇木田建次夏子渊
Owner SICHUAN UNIV
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