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Catalase gene and use thereof

A catalase and amino acid technology, which is applied in the manufacture of alcoholic beverages and in the field of alcoholic beverages, can solve the problems of 1-propanol increase and fermentation delay, etc., and achieve the effect of long shelf life and excellent fragrance stability

Inactive Publication Date: 2007-09-05
SUNTORY HLDG LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition, J.Hansen et al. have made the following attempts to prevent the reduction of generated sulfite by destroying the MET10 gene encoding sulfite reductase (sulfite ionreductase), thereby increasing the amount of sulfite produced (J.Hansen et al. , Nature Biotech., 1587-1591, 1996), but there have also been delays in fermentation and the increase of unwelcome aroma components acetaldehyde and 1-propanol

Method used

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  • Catalase gene and use thereof
  • Catalase gene and use thereof
  • Catalase gene and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0107] Example 1: Cloning of the gene encoding catalase (nonScCTA1) in Saccharomyces cerevisiae

[0108] As a result of searching the comparative database described in JP-A-2004-283169, a gene non-ScCTA1 (sequence number: 1) encoding catalase unique to S. cerevisiae was found. Based on the obtained base sequence information, primers non-ScCTA1_for (sequence number: 5) and non-ScCTA1_rv (sequence number: 6) were designed to amplify the respective full-length genes, and the strain Saccharomycespastorianus Weihenstephan 34 / Chromosomal DNA of 70 strains (sometimes abbreviated as "W34 / 70 strain") was used as a template, and a DNA fragment containing the full-length non-ScCTA1 gene was obtained by PCR.

[0109] The non-ScCTA1 gene fragment obtained above was inserted into pCR2.1-TOPO vector [manufactured by Invitrogen] by TA cloning. The base sequence of the non-ScCTA1 gene was analyzed by the Sanger method (F. Sanger, Science, 214:1215, 1981) to confirm the base sequence.

Embodiment 2

[0110] Example 2: Analysis of non-ScCTA1 gene expression in beer test brewing

[0111] The beer yeast Saccharomyces pastorianus W34 / 70 strain was used for the experimental brewing of beer, and the mRNA extracted from the fermentation yeast cell was detected by the Saccharomyces DNA microarray.

[0112] Wort extract concentration 12.69%

[0113] Wort capacity 70L

[0114] Wort dissolved oxygen concentration 8.6ppm

[0115] Fermentation temperature 15°C

[0116] Yeast input amount 12.8×10 6 cells / mL

[0117]The fermented liquid was sampled over time, and the changes over time in the amount of yeast proliferation (Fig. 1) and apparent extract concentration (Fig. 2) were observed. At the same time, the yeast cells were sampled to prepare mRNA, and the prepared mRNA was biotinylated to be hybridized on the brewer's yeast DNA microarray. Signal detection was performed using GeneChip Operating System [GCOS; GeneChip Operating Software 1.0, manufactured by AFFYMETRIX Corporation...

Embodiment 3

[0118] Example 3: Preparation of non-ScCTA1 gene high expression strain

[0119] The non-ScCTA1 / pCR2.1-TOPO described in Example 1 was digested with restriction enzymes SacI and NotI to prepare a DNA fragment containing the non-ScCTA1 gene. This fragment was ligated with pUP3GLP2 treated with restriction enzymes SacI and NotI to construct a non-ScCTA1 high expression vector pUP-nonScCTA1. pUP3GLP2 refers to the YIp type (chromosomal integration type) yeast expression vector containing the orotidine-5'-phosphate decarboxylase gene URA3 at the homologous recombination site, and the introduced gene is passed through the 3-phosphate glyceraldehyde dehydrogenase gene TDH3 Promoter / terminator high expression. Under the control of the promoter / terminator of the galactokinase gene GAL1, the drug resistance gene YAP1 was introduced as a selectable marker, thereby inducing expression in a galactose-containing medium. Also included as a selectable marker for E. coli ampicillin resistan...

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Abstract

The present invention relates to a gene encoding a catalase and use thereof, in particular, a brewery yeast having high sulfite-producing capability, alcoholic beverages produced with said yeast, and a method for producing said beverages. More particularly, the present invention relates to a yeast, whose capability of producing sulfite, that contribute to stability of flavor in products, is enhanced by amplifying expression level of CTA1 gene encoding a catalase Cta1p, especially non-ScCTA1 gene or ScCTA1 gene specific to a lager brewing yeast, and to a method for producing alcoholic beverages with said yeast.

Description

technical field [0001] The present invention relates to a gene encoding catalase and its use, and in particular to brewer's yeast for producing alcoholic beverages with excellent aroma, alcoholic beverages produced using the yeast, and methods for producing them. More specifically, the present invention relates to improving the expression level of the gene CTA1 encoding catalase Ctalp in Saccharomyces cerevisiae, especially the characteristic nonScCTA1 gene or ScCTA1 gene in Saccharomyces spp. Yeast with improved sulfate production ability, method for producing alcoholic beverages using the yeast, and the like. Background technique [0002] Sulfite is known as a compound with strong antioxidant activity, and is widely used as an antioxidant in the fields of food, beverages, pharmaceuticals, etc. Wait). In alcoholic beverages, sulfites are also used as antioxidants. For example, they play an important role in the quality assurance of wines that need to be aged for a long ti...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/53C12N9/08C12N15/63C12N1/19C12C11/02C12G1/00C12Q1/68
CPCC12N9/0065C12N15/52C12N15/63
Inventor 中尾嘉宏儿玉由纪子下永朋子大村文彦
Owner SUNTORY HLDG LTD
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