Preparation method of yeast fusant being resistant to high salt and being high in yield of ester

A technology of yeast fusion and yeast, which is applied in the direction of microorganism-based methods, biochemical equipment and methods, hybrid cell preparation, etc., can solve the problems of slow growth, unsatisfactory high-quality soy sauce, low ability to produce ester fragrance, etc., and achieve The effect of rapid breeding, no cell wall barrier, and high frequency of gene recombination

Active Publication Date: 2017-07-14
GUANGDONG IND TECHN COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, under high-salt conditions, Torulopsis mogella grows slowly and has a low ability

Method used

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  • Preparation method of yeast fusant being resistant to high salt and being high in yield of ester
  • Preparation method of yeast fusant being resistant to high salt and being high in yield of ester
  • Preparation method of yeast fusant being resistant to high salt and being high in yield of ester

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] (1) Take one ring each of fresh Debaryomyces hansenii ZXL20141126 and Torulopsis mogii ZXL20161207 slant strains, and inoculate them into 250mL shake flasks containing 30mL wort liquid medium , 28°C, 150r / min shaking culture for 12h. 1% (v / v) inoculum amount was transferred to 500mL shake flasks containing 90mL wort liquid culture medium, 28°C, 200r / min shaking culture, Debariae hansenii was cultured for 6h, Mogger Torulopsis globosa was cultured for 8 hours.

[0053] (2) Adjust the concentration of the two kinds of yeast suspensions after activation to 10 7 / mL, take 1mL and add it to a 1.5mL centrifuge tube, centrifuge at 3000r / min for 10min to collect the bacteria; use hypertonic buffer (0.2mol / L KH 2 PO 4 Solution 90.0mL with 0.2mol / LK 2 HPO 4 Mix 10.0 mL of the solution, adjust the pH value to 5.8 with HCl, add 170 g / L sucrose and 0.8 mol / L mannitol respectively) to wash and centrifuge twice, remove the culture medium, and add 3 mL of pretreatment agent (PBS b...

Embodiment 2

[0074] A method for preparing a high-salt-tolerant and high-ester-yielding yeast fusant, comprising the following steps:

[0075] (1) Take one ring each of fresh Debaryomyces hansenii (Debaryomyces hansenii) ZXL20141126 (preservation number GDMCC NO: 60127) and Torulopsis mogii (Torulopsis mogii) ZXL20161207 (preservation number GDMCC NO: 60126) , were inoculated into 250mL shake flasks containing 30mL of wort liquid medium, and cultured with shaking at 28°C and 150r / min for 12h. 1% (v / v) inoculum amount was transferred to 500mL shake flasks containing 90mL wort liquid culture medium, 28°C, 200r / min shaking culture, Debariae hansenii was cultured for 6h, Mogger Torulopsis globosa was cultured for 8 hours.

[0076] (2) Adjust the concentration of the two kinds of yeast suspensions after activation to 10 7 / mL, take 1mL and add it to a 1.5mL centrifuge tube, centrifuge at 3000r / min for 10min to collect the bacteria; use hypertonic buffer (0.2mol / L KH 2 PO 4 Solution 90.0mL w...

Embodiment 3

[0088] (1) Take one ring each of the slant strains of fresh Debaryomyces hansenii ZXL20141126 (preservation number GDMCC NO: 60127) and Torulopsis moggerii ZXL20161207 (preservation number GDMCC NO: 60126), and inoculate them into 30 mL of wort juice respectively. In a 250mL shake flask of liquid medium, shake culture at 28°C and 150r / min for 12h. 1% (v / v) inoculum amount was transferred to 500mL shake flasks containing 90mL wort liquid culture medium, 28°C, 200r / min shaking culture, Debariae hansenii was cultured for 6h, Mogger Torulopsis globosa was cultured for 8 hours.

[0089] (2) Adjust the concentration of the two kinds of yeast suspensions after activation to 10 7 / mL, take 1mL and add it to a 1.5mL centrifuge tube, centrifuge at 3000r / min for 10min to collect the bacteria; use hypertonic buffer (0.2mol / L KH 2 PO 4 Solution 90.0mL with 0.2mol / LK 2 HPO 4 Mix 10.0 mL of the solution, adjust the pH value to 5.8 with HCl, add 170 g / L sucrose, 0.8 mol / L mannitol) to wa...

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Abstract

The invention discloses a preparation method of a yeast fusant being resistant to high salt and being high in yield of ester. The method includes the steps of: 1) inactivating Debaryomyces hansenii protoplast through thermal treatment and Torulopsis mogii protoplast through ultraviolet light; 2) performing electric-fusion to the inactivated Debaryomyces hansenii protoplast and Torulopsis mogii protoplast and culturing the mixture to obtain the fusant; 3) inoculating the fusant on a culture medium, which contains 26% (w/w) of NaCl, to perform cultivation and fermentation, and detecting ester content of a product obtained through fermentation; and 4) detecting growth speed of the fusant to prepare the yeast fusant being resistant to high salt and being high in yield of ester. The method belongs to protoplast electric-fusion yeast breeding methods, is free of barrier of cell walls, is high in fusion frequency of cells and is high in frequency of gene recombination too. The yeast fusant has advantages of parents, is resistant to high salt, is high in yield of ester and is high-temperature resistant.

Description

technical field [0001] The invention relates to the field of yeast breeding, in particular to a method for preparing a high-salt-resistant and high-ester-yielding yeast fusion child. Background technique [0002] In the traditional brewing of soy sauce or miso, there are a variety of salt-tolerant yeasts, and their role in the fermentation process is of great significance. Among them, Torulopsis mogella can impart specific aroma components to soy sauce and sauce, and it breeds in an environment with 18% salt and about 1.4% nitrogen, and is called "osmotic pressure-resistant yeast" or "salt-tolerant yeast". However, Torulopsis moggerii grows slowly under high-salt conditions, and has a low ability to produce ester aroma, which cannot meet the needs of high-quality soy sauce preparation. Therefore, there is an urgent need for a strain that grows faster and has stronger ester-producing ability under high-salt conditions. This is beneficial to improve the ester flavor of soy s...

Claims

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Application Information

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IPC IPC(8): C12N15/04C12N13/00A23L27/50C12R1/645C12R1/88
CPCC12N13/00C12N15/04
Inventor 朱晓立谭才邓庄玉洪张银冰杜淑霞
Owner GUANGDONG IND TECHN COLLEGE
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