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Phosphoadenylyl sulfate reductase gene and use thereof

A technology of phosphoadenosyl sulfate and reductase, which is applied in the field of manufacturing said beverages and alcoholic beverages, and achieves the effects of long shelf life, increased sulfite content, and excellent flavor stability

Inactive Publication Date: 2008-08-20
SUNTORY HLDG LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Donalies et al constructed yeast with high expression of MET16 gene, but the concentration of sulfite generated no matter whether using synthetic medium or wort, no increase (Donalies and Stahl, Yeast, 19, 475-484, 2002: Non-patent literature 2)

Method used

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  • Phosphoadenylyl sulfate reductase gene and use thereof
  • Phosphoadenylyl sulfate reductase gene and use thereof
  • Phosphoadenylyl sulfate reductase gene and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0126] Cloning of Phosphoadenyl Sulfate Reductase Gene (nonScMET16)

[0127] Searching the comparative database described in JP-A-2004-283169 revealed a novel phosphoadenoylsulfate reductase gene nonScMET16 (SEQ ID NO: 1) unique to Saccharomyces cerevisiae. According to the obtained nucleotide sequence information, primers nonScMET16_for (sequence number 3) / nonScMET16_rv (sequence number 4) for amplifying the full-length gene were respectively designed, and the chromosomal DNA of the genome interpretation strain Saccharomyces pastorianus Weihenstephan 34 / 70 was used as The template was subjected to PCR to obtain a DNA fragment (about 0.8 kb) containing the full-length gene of nonScMET16.

[0128] The nonScMET16 gene fragment obtained by the above operation was inserted into pCR2.1-TOPO vector (manufactured by Invitrogen) by TA cloning. The nucleotide sequence of the nonScMET16 gene was analyzed by the Sanger method (F. Sanger, Science, 214, 1215, 1981), and the nucleotide s...

Embodiment 2

[0130] Analysis of nonScMET16 gene expression in beer trial brewing

[0131] Saccharomyces cerevisiae pasteur W34 / 70 strain was used for beer brewing, and the mRNA extracted from cerevisiae cerevisiae cells during fermentation was detected by Saccharomyces cerevisiae DNA microarray.

[0132] Wort extract concentration 12.69%

[0133] Wort volume 70L

[0134] Dissolved oxygen concentration in wort 8.6ppm

[0135] Fermentation temperature 15°C

[0136] The amount of yeast added 12.8×10 6 cells / mL

[0137] The fermentation broth was sampled over time to observe the yeast proliferation ( figure 1 ), the apparent concentration of the extract ( figure 2 ) changes over time. At the same time, the yeast cells were sampled, the prepared mRNA was labeled with biotin, and hybridized on the brewer's yeast DNA microarray. Signal detection was performed with a GeneChip Operating System (GCOS; GeneChip Operating Software 1.0, manufactured by Affymetrix). The expression pattern of...

Embodiment 3

[0139] Production of high expression strain of nonScMET16 gene

[0140] From the plasmid nonScMET16 / pCR2.1-TOPO described in Example 1, a DNA fragment of about 0.8 kb containing the nonScMET16 gene was prepared by treating with restriction enzymes SacI and NotI. It was connected with pUP3GLP2 treated with restriction enzymes SacI and NotI to construct nonScMET16 constitutive expression vector pUP-nonScMET16. The yeast expression vector pUP3GLP2 is a YIp (chromosomally integrated) vector containing the orotidine-5-phosphate decarboxylase gene URA3 at the homologous recombination site, and the introduced gene is in the glyceraldehyde-3-phosphate dehydrogenase gene TDH3 Constitutive expression under the control of the promoter and terminator. As a selection marker in yeast, the drug resistance gene YAP1 was integrated under the control of the promoter and terminator of the galactokinase gene GAL1 to be induced to express in a medium containing galactose. Also contains the amp...

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Abstract

The present invention relates to a phosphate adenylylsulfate reductase gene and uses thereof, and particularly relates to a brewer's yeast for producing an alcoholic beverage with excellent flavor stability, an alcoholic beverage produced using the yeast, a method for producing the beverage, and the like. More specifically, the present invention relates to controlling the expression level of the gene MET16 encoding the phosphate adenylylsulfate reductase Met16p of Saccharomyces cerevisiae, especially the characteristic non-ScMET16 gene of Saccharomyces cerevisiae to control the aroma stability of the product. A yeast capable of producing sulfite, a method for producing an alcoholic beverage using the yeast, and the like.

Description

technical field [0001] The present invention relates to phosphoadenylyl sulfate reductase (phosphoadenylyl sulfate reductase) gene and its use, and in particular to brewing yeast for producing alcoholic beverages with excellent flavor stability, alcoholic beverages produced using the yeast, and methods for producing the beverages, etc. . More specifically, the present invention relates to controlling the expression level of the gene MET16, which is the characteristic non-ScMET16 gene of Saccharomyces cerevisiae, by controlling the expression level of the gene MET16 of the phosphoadenosyl sulfate reductase Met16p of Saccharomyces cerevisiae, which contributes to the stabilization of product aroma. A yeast capable of producing sulfite, a method for producing an alcoholic beverage using the yeast, and the like. Background technique [0002] It is known that sulfite is a compound with strong antioxidant effect, and is widely used as an antioxidant in the fields of food, beverag...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/02C07K14/395C12N15/53C12N15/81C12C11/00C12G1/00C12Q1/68G01N33/573C12R1/86
CPCC12N9/0051C07K14/395C12N15/52C12N9/0004
Inventor 中尾嘉宏儿玉由纪子下永朋子
Owner SUNTORY HLDG LTD
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