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Freezing resistant Saccharomyces cerevisiae bacterial strain and construction method thereof

A technology of baker's yeast and baker's yeast, applied in the field of genetic engineering, can solve the problems of low efficiency of repetitive sequence recombination, and achieve the effect of safe industrial production

Inactive Publication Date: 2013-08-07
TIANJIN UNIVERSITY OF SCIENCE AND TECHNOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, in this method, the 40bp repeat sequence recombination efficiency is low

Method used

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  • Freezing resistant Saccharomyces cerevisiae bacterial strain and construction method thereof
  • Freezing resistant Saccharomyces cerevisiae bacterial strain and construction method thereof
  • Freezing resistant Saccharomyces cerevisiae bacterial strain and construction method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1: Construction of baker's yeast with seamless knockout of neutral trehalase gene NTH1

[0024] The starting strain CICC31616 used in this example, the transformed baker's yeast strain (Saccharomyces cerevisiae) in this example was preserved on March 28, 2013 in the General Microorganism Center (CGMCC) of China Microbiological Culture Management Committee, and the preservation number is CGMCC No.7379 . The source of the plasmid YIplac211 can be found in R.D.Gietz, and A.Sugino, New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites. Gene 74 (1988) 527-34. The Escherichia coli DH5a was purchased from Takara Company. The YPD medium is a universal complete medium, the yeast synthetic medium (SD) consists of 2% glucose, 0.67% YNB, and an amino acid mixture solution without uracil, and the solid medium contains 2% imported agar powder.

[0025] According to the yeast genome data in Geneba...

Embodiment 2

[0041] Example 2: Comparison of intracellular trehalose accumulation in freeze-resistant baker's yeast and determination of cell survival rate after freezing

[0042] The haploid parents and their corresponding recombinant strains were cultured for 48 hours at the same time, and the intracellular trehalose accumulation level and the cell survival rate 21 days after freezing the liquid simulated dough were measured. The results are shown in Table 2. As shown in the results, after 48 hours of cell culture, the intracellular trehalose of the constructed strain accounted for 11.7% of the dry cell weight, which was 134% higher than that of the parental strain (5.0%); After culturing, it was frozen for 21 days, and the cell survival rate was measured. The cell survival rate of the mutant strain was 78%, which was 2.6 times that of the starting strain (30%).

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Abstract

The invention discloses a freezing resistant Saccharomyces cerevisiae bacterial strain and construction method thereof, concretely is a freezing resistant Saccharomyces cerevisiae BY-14A delta U constructed by a non-trace knock-out gene NTH1, and the preservation number is CGMCC NO.7379. The freezing resistant Saccharomyces cerevisiae constructed by a non-trace gene knock-out method is realized by fusing PCR and a yeast integrative plasmid YIplac211, and knocking out a gene NTH1 for coding neutral trehalase. The invention realizes a rapid and high efficiency non-trace knock out of Saccharomyces cerevisiae NTH1 gene, and intracellular mycose content increases by 134% than that of the parent bacterial strain without influences to other fermentation performances of the bacterial strain BY-14A delta U; after the cells in the liquid dough are frozen for 21 days, the survival rate reaches to 78%, which is 2.6 times of the parent bacterial strain. The genome in the constructed microzyme does not have any residual exogenous genes, therefore the product can be used for industrial production safely.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a freeze-resistant baker's yeast strain and a gene traceless knockout method thereof. Background technique [0002] Baker's Yeast (Baker's Yeast, scientific name Saccharomyces cerevisiae) is an important industrial microorganism, rich in nutrients, and is the most important microbial leavening agent, bioloosening agent and bionutrient in the production of steamed bread, bread, biscuits and other pasta. With the rapid development of frozen dough production technology, freeze-resistant baker's yeast and its freeze-resistant mechanism have been extensively studied. In developed countries such as Europe, America, and Japan, frozen dough technology has also been widely used in the baking industry. The poor freezing resistance of common baker's yeast is the main factor limiting the development of frozen dough technology. Trehalose is a self-protective agent in yeast cells, which can...

Claims

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Application Information

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IPC IPC(8): C12N15/09C12R1/865C12N1/18
Inventor 肖冬光王光路董建张翠英
Owner TIANJIN UNIVERSITY OF SCIENCE AND TECHNOLOGY
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