Multiple primers, kit and detection method for detecting fermentation conditions affecting the expression level of Saccharomyces cerevisiae protease a gene

A technology of gene expression and fermentation conditions, applied in the field of multiple primers of fermentation conditions, can solve the problems of long detection cycle, time-consuming, cumbersome detection process, etc., to improve beer quality, ensure reliability and repeatability, and strong specificity. sexual effect

Active Publication Date: 2021-08-10
TSINGTAO BREWERY
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Problems solved by technology

[0004] This application aims at the problems of hysteresis, time-consuming, cumbersome detection process and long detection period when detecting the fermentation conditions affecting the expression of beer yeast protease A gene by measuring the activity of protease A. Multiple primers, kit and screening method for fermentation conditions of gene expression

Method used

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  • Multiple primers, kit and detection method for detecting fermentation conditions affecting the expression level of Saccharomyces cerevisiae protease a gene
  • Multiple primers, kit and detection method for detecting fermentation conditions affecting the expression level of Saccharomyces cerevisiae protease a gene
  • Multiple primers, kit and detection method for detecting fermentation conditions affecting the expression level of Saccharomyces cerevisiae protease a gene

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Experimental program
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Embodiment 1

[0026] A method for rapid detection of industrial Pasteurella protease A gene, comprising extraction of total RNA, reverse transcription reaction to prepare cDNA template, multiple PCR reaction, capillary electrophoresis, and product fragment analysis.

[0027] (1) Multiple primer design

[0028] According to the full-length sequences of PEP4-Sc and PEP4-Sb of the protease A gene PEP4-Sc and PEP4-Sb of P. industrialis and internal reference genes ACT1-Sc and ACT1-Sb, specific upstream and downstream multiple primers suitable for GeXP detection were designed (Table 1).

[0029] Table 1 The specific upstream and downstream primers of the industrial Pasteurella protease A gene and internal reference genes

[0030]

[0031] (2) Total RNA extraction and purification

[0032] Follow the procedure of the RNA extraction kit and operate step by step to extract yeast RNA.

[0033] Add 200 μL of phosphate buffer to the sample, centrifuge at 12000 rpm, and remove the supernatant. Ad...

Embodiment 2

[0059] Get the wort of A factory and B factory and the corresponding fermented liquid of the same period, analyze the content of glutamic acid and histidine in the wort, record the situation of yeast vigor, yeast death rate and yeast algebra, measure with the method described in Example 1 Expression level of protease A gene of yeast in fermentation broth. The experimental results are shown in Table 3:

[0060] The protease A gene expression level result of yeast in the fermented liquid of factory A and B factory of table 3

[0061]

[0062] From the above experimental results, it can be found that by adjusting the wort formula and the parameters of the beer fermentation process, the expression of protease A gene of yeast in the fermentation broth can be effectively reduced, thereby improving the beer foam retention and improving the quality of beer. Simultaneously, the multiple primers of the present invention are used to detect the expression of yeast protease A gene, whi...

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Abstract

The invention proposes a multiple primer, a kit and a detection method for detecting fermentation conditions that affect the expression of beer yeast protease A gene, which belong to the field of food detection and can solve the traditional problem of detecting protease A that affects beer yeast protease A by measuring the activity of protease A. The fermentation conditions of gene expression amount have the problems of long time-consuming, cumbersome detection process and long detection cycle. The technical scheme includes the design of multiple primers, reverse transcription and synthetic cDNA template using the total RNA of yeast strains extracted under different fermentation conditions as a template, multiple PCR amplification reactions, capillary electrophoresis separation, and the use of GeXP system for capillary electrophoresis separation. The results were analyzed; using the internal reference gene as a control, the expression level of the protease A gene was calculated in combination with the peak area of ​​the corresponding peak of the key gene, and the fermentation conditions affecting the expression level of the protease A gene in S. cerevisiae were judged according to the expression level of the protease A gene.

Description

technical field [0001] The invention relates to the field of food detection, in particular to multiple primers, a kit and a screening method for detecting fermentation conditions affecting the expression of brewer's yeast protease A gene. Background technique [0002] White, delicate and long-lasting foam is one of the important symbols of high-quality beer. The persistence of foam is measured by the index of foam retention. The national standard stipulates the standard of foam retention of premium beer, bottled wine ≥ 180 seconds, canned wine ≥ 150 seconds. Protein is the skeleton component of beer foam, and the attenuation of foam retention is mainly due to the destruction of protein by protease A. Protease A is derived from brewer's yeast. It will be released when the yeast is autolyzed, and gradually decomposes the hydrophobic protein in the fermentation broth or changes the characteristics of the hydrophobic protein, thereby affecting the foam retention of the fermenta...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6851C12N15/11
CPCC12Q1/6851C12Q2537/143C12Q2531/113
Inventor 董建军余俊红尹花黄淑霞胡淑敏贺扬刘佳马增新
Owner TSINGTAO BREWERY
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