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Brewer's yeast engineering bacterium producing 2-phenylethyl alcohol, preparation method and applications thereof

A technology of Saccharomyces cerevisiae and engineering bacteria, applied in the field of genetic engineering, can solve the problems of limited 2-phenylethanol synthesis ability and difficulty in large-scale industrial production, and achieve the effects of improved synthesis ability, short fermentation cycle and simple fermentation conditions

Active Publication Date: 2016-06-08
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Existing research mainly focuses on natural screening of strains, traditional microbial breeding or improvement of fermentation process, but there is a problem that the synthesis capacity of 2-phenylethanol is limited and it is difficult to carry out large-scale industrial production

Method used

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  • Brewer's yeast engineering bacterium producing 2-phenylethyl alcohol, preparation method and applications thereof
  • Brewer's yeast engineering bacterium producing 2-phenylethyl alcohol, preparation method and applications thereof
  • Brewer's yeast engineering bacterium producing 2-phenylethyl alcohol, preparation method and applications thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Embodiment 1, the construction of the Saccharomyces cerevisiae engineered bacteria that produces 2-phenylethanol

[0041] 1. Acquisition of GATA transcription factor Gln3p encoding gene GLN3

[0042] (1) PCR amplification of the gene GLN3 encoding the GATA transcription factor Gln3p

[0043] According to the GLN3 gene sequence (GenBank number is M35267) of the coding GATA transcription factor Gln3p of the reported Saccharomyces cerevisiae, design following primers:

[0044] Upstream primer P1: 5′-ACGC GTC GACAACAAATGCAAGACGACC-3' (SEQ ID No. 1)

[0045] (The underlined sequence is the SalI restriction recognition site)

[0046] Downstream primer P2: 5′-ACGC GAGCTC AATACGCGGTCATATACC-3' (SEQ ID No. 2)

[0047] (The underlined sequence is the SacI restriction recognition site)

[0048] (2) Using the genomic DNA of Saccharomyces cerevisiae MT2 with a higher yield of 2-phenylethanol as a template, and using P1 and P2 as primers to perform PCR amplification to obtain...

Embodiment 2

[0117] Embodiment 2, yeast fermentation method produces 2-phenylethanol

[0118] 1. Transfer the activated MT2 (YEpKAG), MT2 (YEpKA) and MT2 into a 250 mL Erlenmeyer flask containing 50 mL of fermentation medium, and culture on a shaker at 30°C and 220 rpm for 36 hours.

[0119] Above-mentioned fermentation medium is made up of solvent and solute, and solvent is water, and solute is L-phenylalanine, glucose, magnesium sulfate, potassium dihydrogen phosphate; The concentration of described L-phenylalanine in fermentation medium is 5g / L, the concentration of the glucose in the fermentation medium is 30g / L, the concentration of the magnesium sulfate in the fermentation medium is 0.5g / L, and the concentration of the potassium dihydrogen phosphate in the fermentation medium is 0.5 g / L.

[0120] 2. After the fermentation finishes, take MT2 (YEpKAG), MT2 (YEpKA) and MT2 fermentation broth, centrifuge to get the supernatant, and measure the 2-phenylethanol output in the fermentation...

Embodiment 3

[0127] Embodiment 3, yeast fermentation method produces 2-phenylethanol

[0128] 1. Transfer the activated MT2 (YEpKAG), MT2 (YEpKA) and MT2 into a 250 mL Erlenmeyer flask containing 50 mL of fermentation medium, and culture on a shaker at 30°C and 220 rpm for 36 hours.

[0129] Above-mentioned fermentation medium is made up of solvent and solute, and solvent is water, and solute is L-phenylalanine, glucose, magnesium sulfate, potassium dihydrogen phosphate; The concentration of described L-phenylalanine in fermentation medium is 5g / L, the concentration of the glucose in the fermentation medium is 40g / L, the concentration of the magnesium sulfate in the fermentation medium is 0.5g / L, and the concentration of the potassium dihydrogen phosphate in the fermentation medium is 0.5 g / L.

[0130] 2. After the fermentation finishes, take MT2 (YEpKAG), MT2 (YEpKA) and MT2 fermentation broth, centrifuge to get the supernatant, and measure the 2-phenylethanol output in the fermentation...

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Abstract

The invention discloses a brewer's yeast engineering bacterium producing 2-phenylethyl alcohol, a preparation method and applications thereof. The invention discloses a preparation method of a brewer's yeast engineering bacterium producing 2-phenylethyl alcohol. According to the preparation method, an encoding gene of GATA transcription factor Gln3p is introduced into an acceptor (brewer's yeast) to obtain the brewer's yeast engineering bacterium producing 2-phenylethyl alcohol. Compared with the host bacterium, the substrate conversion rate and cell's 2-phenylethyl alcohol synthesizing performance of brewer's yeast engineering bacterium are prominently improved; moreover, the fermentation conditions of the engineering bacterium are simple, the fermentation period is short, and the engineering bacterium can be applied to industrial production of 2-phenylethyl alcohol.

Description

technical field [0001] The invention relates to a Saccharomyces cerevisiae engineering bacterium producing 2-phenylethanol and a preparation method and application thereof, belonging to the field of genetic engineering. Background technique [0002] 2-phenylethanol (2-phenylethanol) is an aromatic alcohol with a rose flavor, which exists in various plant essential oils such as rose and jasmine, and is also a natural flavor substance of various fermented foods such as wine, rice wine, beer, and bread. A key factor in determining the quality of fermented foods. The aroma of 2-phenylethyl alcohol is quite popular, and it is the mainstream style of international flavors and fragrances. It is widely used in the fields of food and daily chemical products. Its usage is second only to vanillin, and it is the second largest fragrance ingredient. In addition, 2-phenylethanol, as a fungicide and angiotensin-converting enzyme inhibitor, also has important applications in the pharmaceut...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/81C12N1/19C12P7/22C12R1/865
Inventor 陈先锐王肇悦白雪晶郭雪娜何秀萍张博润
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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