Xylose fermentation strain resistant to inhibitors and construction method and applications thereof

A fermentation strain and construction method technology, applied in the field of yeast, can solve problems such as deletion, complex construction steps of Saccharomyces cerevisiae strain, etc., and achieve improved inhibitor tolerance, superior inhibitor tolerance, and increased xylose consumption rate Effect

Active Publication Date: 2019-05-03
CHINA PETROLEUM & CHEM CORP +2
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  • Description
  • Claims
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Problems solved by technology

[0003] Chinese patent CN105985915A discloses a genetically engineered strain in which a GRE3 gene having the nucleotide sequence shown in SEQ ID NO: 1 is deleted in Saccharomyces cerevisiae 1308-P. The construction method of the engineered strain is: GRE3 gene knockout component 1. Transfer the GRE3 gene knockout component into yeast, knock out the GRE3 gene\remove t...

Method used

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  • Xylose fermentation strain resistant to inhibitors and construction method and applications thereof
  • Xylose fermentation strain resistant to inhibitors and construction method and applications thereof
  • Xylose fermentation strain resistant to inhibitors and construction method and applications thereof

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Embodiment 1

[0040] This embodiment is the construction method of the xylose fermentation strain SEB11 that is tolerant to inhibitors. The starting strain used in this embodiment is Saccharomyces cerevisiae SEB5 (CGMCC No.11325, General Microbiology Center of China Committee for Culture Collection of Microorganisms), which has Excellent performance of fermenting xylose to produce ethanol. The construction method includes the following steps:

[0041] (1) plasma mutagenesis treatment;

[0042]Select a loop of SEB5 strain into 2% YPD liquid medium, put it into a 30°C constant temperature shaker at 160rpm for pre-cultivation. After 16 hours of pre-cultivation, 1 mL of the bacterial liquid was taken, centrifuged at 10,000 rpm for 3 minutes, the bacterial cells were collected, transferred to 2% YPD, and placed in a constant temperature shaker at 160 rpm at 30°C. When cultured for 8 hours, the starting yeast strain SEB5 was in the transition period between the logarithmic growth phase and the ...

Embodiment 2

[0047] The present embodiment is a fermentation experiment in which xylose is the sole carbon source under the condition of acetic acid (80mmol / L), which adopts the following steps:

[0048] SEB11 was inoculated on 2% YPD solid plates and activated in a 30°C incubator for 24 hours. A ring of activated yeast was selected and inoculated into 100 mL of 5% YPD liquid medium, and incubated at a constant temperature for 16 hours (30° C., 160 rpm). The pre-cultured bacterial liquid was centrifuged to collect the thalline, inoculated 2.5g of wet thallus into 100mL of 5% YPX medium containing 80mmol / L acetic acid, and fermented in a constant temperature water bath (200rpm, 35°C). The results were as follows: figure 1 shown.

[0049] figure 1 Part A is the fermentation result of SEB11, figure 1 Part B of is the fermentation result of the starting strain SEB5. Compared with the original strain, the xylose consumption rate of SEB11 was significantly increased. The consumption rate of ...

Embodiment 3

[0051] The present embodiment is a fermentation experiment in which xylose is the only carbon source under the condition of the presence of 5-HMF (15mmol / L), and it adopts the following steps:

[0052] SEB11 was inoculated on 2% YPD solid plates and activated in a 30°C incubator for 24h. A ring of activated yeast was selected and inoculated into 100 mL of 5% YPD liquid medium, and incubated at a constant temperature for 16 hours (30° C., 160 rpm). The pre-cultured bacterial liquid was centrifuged to collect the bacterial cells, inoculated 2.5g of wet bacterial cells into 100mL of 5% YPX medium containing 15mmol / L 5-HMF, and fermented in a constant temperature water bath (200rpm, 35°C), the results Such as figure 2 shown.

[0053] figure 2 Part A is the fermentation result of SEB11 under this condition, figure 2 Part B is the fermentation result of the starting strain SEB5 under this condition. 15mmol / L 5-HMF had no obvious inhibitory effect on SEB11, showing better tol...

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Abstract

The invention discloses a xylose fermentation strain resistant to inhibitors. The strain is named as SEB11 and has a preservation number of CGMCC No. 16570. The invention also relates to a construction method of the above-mentioned xylose fermentation strain, and the method comprises the steps of: selecting a starting strain SEB5; performing plasma mutagenesis treatment on the starting strain, andpicking out bacterial colonies which are better or similar to the starting strain in growth situation; and subjecting the picked strains to plate initial screening and actual saccharification liquidrescreening so as to obtain the xylose fermentation strain SEB11 resistant to inhibitors. The present invention also relates to applications of the above xylose fermentation strain in producing ethanol by using xylose in lignocellulose. The method adopts an industrial strain of the xylose as a starting strain, adopts a normal-pressure room-temperature plasma mutagenesis technique for mutagenesis,and obtains the xylose fermentation mutant strain SEB11 with excellent inhibitor tolerance according to the growth condition in a plate medium containing inhibitors and the actual saccharification liquid fermentation rescreening. The xylose consumption rate is significantly increased, and a better fermentation result exhibits.

Description

technical field [0001] The invention relates to a yeast, in particular to a xylose fermentation strain resistant to inhibitors and its construction method and application. Background technique [0002] With the depletion of fossil resources such as petroleum and the intensification of the greenhouse effect, it has become the focus of people's attention to find alternatives to petroleum resources. Using lignocellulose to produce fuel ethanol to replace fossil fuels can effectively solve the energy crisis and environmental pollution caused by fossil fuels. However, several obstacles exist in the conversion of lignocellulosic ethanol, including inhibitor tolerance and low xylose utilization. Inhibitors formed by acid-catalyzed hydrolysis of lignocellulose (mainly including furan derivatives, weak acids, and phenolic compounds) reduce the growth rate and fermentation of S. cerevisiae, thereby reducing ethanol production. Although there are many reports on the ability of Saccha...

Claims

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Application Information

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IPC IPC(8): C12N1/18C12N15/01C12N13/00C12P7/10C12R1/865
CPCY02E50/10
Inventor 缪晡程华陈栋汤岳琴吴娅箐丁伟军
Owner CHINA PETROLEUM & CHEM CORP
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