CN115786576A - Malt PYF performance prediction method based on barley gene expression in malting process and application thereof - Google Patents
Malt PYF performance prediction method based on barley gene expression in malting process and application thereof Download PDFInfo
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Abstract
The invention provides a malt PYF performance prediction method based on barley gene expression in a malting process and application thereof, belongs to the field of biology, and can solve the technical problems of complex operation steps, long time consumption, narrow application range and the like of a traditional malt PYF detection method. The prediction method specifically comprises the following steps: screening candidate genes, screening tracking genes, carrying out reverse transcription reaction, designing primers for multiple PCR reactions, extracting RNA, preparing a cDNA template by reverse transcription, carrying out multiple PCR reactions, carrying out electrophoresis and expression quantity analysis on PCR amplification products, predicting the PYF performance of malt and the like. The method selects the tracking gene by comparing the gene expression difference of the standard malt and the PYF malt in the malting process and combining the large production process, and finally realizes the quick and accurate analysis of the PYF performance of the malt by comparing the expression quantity conditions of the standard malt and the malt to be detected and combining the evaluation standard. The method can be applied to the prediction of the PYF performance of the malt.
Description
Technical Field
The invention belongs to the field of bioengineering, and particularly relates to a malt PYF performance prediction method based on barley gene expression in a malting process and application thereof.
Background
Pre-Flocculation (PYF) of Yeast refers to the phenomenon that Yeast flocculates and settles when sufficient fermentation time is not reached in the beer fermentation stage. The phenomenon can cause slow sugar degradation and slow acetaldehyde reduction in the fermentation process, so that the maturity of the finished beer is poor, the fermentation quality of the beer is influenced, and the quality problem of the beer is caused. The PYF problem is a quality problem specific to the beer industry, due to the presence of factor substances in the malt that cause the yeast to flocculate prematurely. Therefore, the detection of the PYF performance of the malt is a detection index which is commonly prepared in the beer industry.
At present, the PYF of malt is generally detected by preparing malt into wort, adding yeast for fermentation, and then detecting the number of suspended yeast in the fermentation liquor. The method needs 40-48 hours of fermentation time, and the total detection time of 3-4 days is needed by adding the detection of the prepared wort and the yeast number, so the detection time is longer. Meanwhile, the method detects that the finished malt is lack of prejudgement to the malt production process, and cannot adjust the malt production process in time.
Therefore, how to develop the malt PYF performance prediction method which can be analyzed in the malt production process, is short in time consumption, high in detection efficiency and high in accuracy has positive significance for guiding the adjustment of the malting process and improving the malt quality.
Disclosure of Invention
Aiming at the technical problems that the conventional malt PYF detection method is complicated in operation steps and long in time consumption, can only detect finished malt and is narrow in application range, the invention provides the PYF performance prediction method which can be analyzed in the malt production process and has the characteristics of short time consumption, high detection efficiency, high accuracy and the like.
In order to achieve the purpose, the invention adopts the technical scheme that:
the malt PYF performance prediction method based on barley gene expression in the malting process comprises the following steps:
analyzing the gene expression difference of the standard malt and the PYF malt in the malting process through a transcriptome, and selecting a plurality of genes with larger expression difference as candidate genes;
tracking the mass production process of the standard malt and the PYF malt, and further selecting the candidate genes with the most obvious variation trend as tracking genes;
designing a reverse transcription reaction primer group and a multiple PCR reaction primer group according to the tracked gene sequence;
extracting total RNA of the malt to be detected and the standard malt;
taking the extracted total RNA as a template, and taking the reverse transcription reaction primer group as a primer to carry out reverse transcription reaction to prepare a cDNA template;
and (3) performing multiple PCR reaction by taking the reverse transcription reaction product as a template and the multiple PCR reaction primer group as a primer, and performing electrophoresis and expression analysis on the multiple PCR reaction products of the malt to be detected and the standard malt to predict the PYF performance of the malt.
In one embodiment, the candidate gene is selected by transcriptionally analyzing differences in gene expression during germination of the standard malt and the PYF malt during malting.
In one embodiment, the screening criteria for the tracking gene is:
the variation amplitude of gene expression is more than or equal to 2.0 or less than 0.5;
wherein the gene expression variation amplitude is the ratio of the PYF malt gene expression principal component score to the standard malt gene expression principal component score.
In one embodiment, the tracking genes include 13 genes including AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17, and WRKY 5.
In one embodiment, the reverse transcription reaction primer set is designed to obtain 13 primers aiming at 13 tracking genes AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17 and WRKY5, and the primers are respectively shown in SEQ ID NO.2, SEQ ID NO.4, SEQ ID NO.6, SEQ ID NO.8, SEQ ID NO.10, SEQ ID NO.12, SEQ ID NO.14, SEQ ID NO.16, SEQ ID NO.18, SEQ ID NO.20, SEQ ID NO.22, SEQ ID NO.24 and SEQ ID NO. 26.
In one embodiment, the multiplex PCR primer set is designed to obtain 13 primers for 13 tracking genes AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17 and WRKY5, which are respectively shown in SEQ ID NO.1, SEQ ID NO.3, SEQ ID NO.5, SEQ ID NO.7, SEQ ID NO.9, SEQ ID NO.11, SEQ ID NO.13, SEQ ID NO.15, SEQ ID NO.17, SEQ ID NO.19, SEQ ID NO.21, SEQ ID NO.23 and SEQ ID NO. 25.
In one embodiment, the malt PYF performance is assessed by performing electrophoresis and expression analysis on the multiple PCR reaction products of the test malt and the standard malt using the following criteria:
obtaining the principal component scores of the standard malt and the malt to be detected through principal component analysis, and taking the absolute value of the difference between the two scores;
when the absolute value of the difference value of the two is less than 0.5, the PYF value of the malt to be detected is more than or equal to 90 percent;
when the absolute value of the difference value is more than 0.5 and less than 5.0, the value of the PYF of the malt to be detected is 50-90 percent;
when the absolute value of the difference value of the two is more than 5.0, the PYF value of the malt to be detected is less than 50 percent.
In one embodiment, the malt PYF performance criterion is obtained by:
performing gene expression analysis on the tracking genes of different PYF malts to obtain a gene expression principal component score;
comparing the gene expression principal component score to a gene expression principal component score of a standard malt, thereby establishing the malt PYF performance criterion.
The invention also provides an application of the malt PYF performance prediction method based on barley gene expression in the malting process in the malt PYF performance prediction.
In one embodiment, the malt comprises finished malt, barley, and malted barley malt germinated in a malting process.
Compared with the prior art, the invention has the advantages and positive effects that:
1. the invention provides a malt PYF performance prediction method based on barley gene expression in the malting process, which selects a tracking gene by comparing the gene expression difference of standard malt and PYF malt in the malting process and combining with the large production process, and realizes accurate prediction of the malt PYF performance by comparing the expression quantity conditions of the standard malt and the malt to be detected and combining with an evaluation standard;
2. according to the malt PYF performance prediction method based on barley gene expression in the malting process, in order to improve the accuracy of a prediction result, a dual standard including a tracking gene screening standard and a malt PYF performance judgment standard is established, and the rapid and accurate analysis of the malt PYF performance can be realized in the malt production process;
3. the malt PYF performance prediction method based on barley gene expression in the malting process can predict the condition of the malt PYF in advance in the malting stage, and has certain guiding significance for actual production.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The embodiment of the invention provides a malt PYF performance prediction method based on barley gene expression in a malting process, which comprises the following steps:
s1, analyzing the gene expression difference of standard malt and PYF malt in the malting process through a transcriptome, and selecting a plurality of genes with larger expression difference as candidate genes;
s2, tracking the large production process of the standard malt and the PYF malt, and further selecting the candidate genes with the most obvious change trend as tracking genes;
s3, designing a reverse transcription reaction primer group and a multiple PCR reaction primer group according to the tracked gene sequence;
s4, extracting total RNA of the malt to be detected and the standard malt;
s5, performing reverse transcription reaction by taking the extracted total RNA as a template and a reverse transcription reaction primer group as a primer to prepare a cDNA template;
and S6, performing multiple PCR reaction by using the reverse transcription reaction product as a template and the multiple PCR reaction primer group as a primer, and performing electrophoresis and expression analysis on multiple PCR reaction products of the malt to be detected and the standard malt to predict the PYF performance of the malt.
In one embodiment, the candidate gene is screened by transcriptionally analyzing the difference in gene expression between the standard malt and the PYF malt during malting, particularly during germination.
In the embodiment, the method for predicting the PYF performance of the malt based on the barley gene expression in the malting process is characterized in that a tracking gene is selected by comparing the gene expression difference of the standard malt and the PYF malt in the malting process and combining the large production process, and then the accurate prediction of the PYF performance of the malt is realized by comparing the expression quantity conditions of the standard malt and the malt to be tested and combining the evaluation standard.
In one embodiment, the screening criteria for the tracking gene are:
the variation amplitude of gene expression is more than or equal to 2.0 or less than 0.5;
wherein the variation range of the gene expression is the ratio of the PYF malt gene expression principal component score to the standard malt gene expression principal component score.
In the above embodiment, the purpose of setting the screening criteria for the trace genes to a range in which the expression of the genes varies by 2.0 or more and less than 0.5 is mainly to find genes whose expression varies greatly before and after the gene expression as characteristic genes. It is generally considered that the difference in expression amounts of 2-fold or more is large. The ratio may be increased to two or more times or decreased to 1/2 or less of the original ratio, and therefore, the ratio is set to 2.0 or more and 0.5 or less.
In one embodiment, the tracking genes include 13 genes including AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17, and WRKY 5.
The functions of the 13 genes mentioned in the above embodiments are as follows:
in a specific embodiment, the reverse transcription reaction primer group is designed to obtain 13 primers aiming at 13 tracking genes AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17 and WRKY5, and the primers are respectively shown as SEQ ID NO.2, SEQ ID NO.4, SEQ ID NO.6, SEQ ID NO.8, SEQ ID NO.10, SEQ ID NO.12, SEQ ID NO.14, SEQ ID NO.16, SEQ ID NO.18, SEQ ID NO.20, SEQ ID NO.22, SEQ ID NO.24 and SEQ ID NO. 26.
In a specific embodiment, the multiple PCR reaction primer set is designed to obtain 13 primers aiming at 13 tracking genes AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17 and WRKY5, and the primers are respectively shown in SEQ ID NO.1, SEQ ID NO.3, SEQ ID NO.5, SEQ ID NO.7, SEQ ID NO.9, SEQ ID NO.11, SEQ ID NO.13, SEQ ID NO.15, SEQ ID NO.17, SEQ ID NO.19, SEQ ID NO.21, SEQ ID NO.23 and SEQ ID NO. 25.
In one embodiment, after performing electrophoresis and expression analysis on multiple PCR reaction products of the malt to be tested and the standard malt, the performance of the malt PYF is predicted by the following method:
(1) Obtaining the principal component scores of the standard malt and the malt to be detected through principal component analysis, and taking the absolute value of the difference between the two scores;
when the absolute value of the difference value of the two is less than 0.5, the PYF value of the malt to be detected is more than or equal to 90 percent;
when the absolute value of the difference value of the two is more than 0.5 and less than 5.0, the PYF value of the malt to be detected is 50-90 percent;
when the absolute value of the difference value of the two is more than 5.0, the PYF value of the malt to be detected is less than 50 percent.
In one embodiment, the malt PYF performance criterion is obtained by:
(1) Performing gene expression analysis on the tracking genes of different PYF malts to obtain a gene expression principal component score;
the different PYF malts mentioned in the above step (1) refer to malts with different PYF values (specifically PYF-90% malt, PYF-50% malt and PYF-30% malt are included), and by analyzing the gene expression of the malts with different PYF values, the expression principal component score and the judgment standard based on the score are obtained with higher accuracy.
Further, the reason why the gene expression principal component score is selected as the analysis index in the present invention is that: in order to comprehensively evaluate the expression of a plurality of genes, the dimension reduction processing is performed on a plurality of gene expression data by using a principal component analysis method, so that the comprehensive quantitative score of a plurality of genes can be obtained.
(2) The gene expression principal component score is compared with the gene expression principal component score of a standard malt, thereby establishing a malt PYF performance criterion.
The invention also provides an application of the malt PYF performance prediction method based on barley gene expression in the malting process in the malt PYF performance prediction.
In a specific embodiment, the malt includes, but is not limited to, finished malt, barley, and malted barley malt germinated in a malting process.
In order to more clearly and specifically describe the method for predicting the PYF performance of malt based on barley gene expression in the malting process and the application thereof, the embodiment of the present invention will be described with reference to the following specific examples.
Example 1
The embodiment provides a screening process of candidate genes and tracking genes in a malt PYF performance prediction method based on barley gene expression in a malting process, which specifically comprises the following steps:
(1) Researches show that the problem of PYF generation by malt is related to stress reaction of barley under external stimulation in the planting and germination processes, so that the invention analyzes the gene expression difference of the standard malt and the PYF malt in the malting process, particularly in the seed germination process through transcriptome analysis, and discovers a plurality of genes with larger expression difference as candidate genes (about 13);
(2) Combining the results of the step (1), selecting 13 genes with the most obvious change trend from the green malt as the tracking genes by tracking the large production process (namely, taking the green malt after the wheat soaking in the wheat preparation process, wherein the RNA extraction should be immediately carried out on the taken sample or the sample is placed at minus 80 ℃ for storage in order to ensure that the life activity of the green malt is not changed), wherein the screening standard of the tracking genes is as follows: the variation amplitude of gene expression is more than or equal to 2.0 or less than 0.5, and the variation amplitude of gene expression is the ratio of PYF malt gene amount to standard malt gene expression amount (the related data are shown in Table 1);
TABLE 1 statistics of Gene expression in Standard malt and PYF malt
Example 2
The embodiment provides a determining process of a malt PYF performance judgment standard in a malt PYF performance prediction method based on barley gene expression in a malting process, which specifically comprises the following steps:
the gene expression of 3 different PYF malts (PYF-90% malt, PYF-50% malt and PYF-30% malt) was analyzed and tested to obtain the score of gene expression principal component, and the relative standard was determined by comparing with the score of standard malt (see Table 2 for the relevant data).
TABLE 2 score of the principal Components of Standard malt and PYF malt
As can be seen from the above data, when the gene expression principal component scores of the malts with different PYF values are compared with those of the standard malt, the absolute value of the difference between the PYF-90% malt gene expression principal component score and the standard malt principal component score is less than 0.5, the absolute value of the difference between the PYF-50% malt gene expression principal component score and the standard malt principal component score is more than 0.5, and less than 5, and the absolute value of the difference between the PYF-30% malt gene expression principal component score and the standard malt principal component score is more than 5.
Example 3
The embodiment provides a method for predicting the PYF performance of malt based on barley gene expression in the malting process, and the embodiment predicts the PYF performance of a newly purchased batch A of barley, and specifically comprises the following steps:
(1) Monitoring gene selection: selecting 13 genes including AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17 and WRKY5 as monitoring genes;
(2) Sampling a sample: during the process of malting, taking green malt after malting (note: in order to ensure that the life activity of the green malt is not changed, the taken sample should be immediately subjected to RNA extraction, or placed at-80 ℃ for storage);
(3) Green malt crushing: in order to ensure that RNA in the green malt is not degraded, the green malt is ground and crushed by liquid nitrogen;
(4) RNA extraction:
extracting RNA from the crushed green malt by adding Trizol (note: trizol is a total RNA extraction reagent, and can directly extract total RNA from cells or tissues);
taking 50-100mg of sample, adding 1ml of Trizol, shaking for lysis, centrifuging at 12000g at 4 ℃ for 5min, transferring the supernatant to a Phasemarker tube, and standing for 5min; adding 0.2ml chloroform, manually shaking for 15s, and standing for 10min; centrifuging at 12000g at 4 deg.C for 10min; taking supernatant (450-550 microliters) to an EP tube, adding 250 microliters of 96% ethanol, and mixing by using a gun tip; transferring the mixed solution onto a silica gel tube, centrifuging at 12000g for 1min, and removing the waste liquid; 700. Mu.l of WB1 was added thereto and 12000g was centrifuged for 1min to remove the waste liquid flowing down. Adding 500 microliter WB2 and 12000g, centrifuging for 1min, and removing the flowing waste liquid; adding 500 microliter WB2, centrifuging at 12000g for 1min, and transferring the silicone tube to a 1.5ml EP tube; adding 50 microliters of clean-free water, standing for 1min, and centrifuging for 1min at 12000g; collecting the centrifuged RNA solution, and storing at-80 ℃;
(5) RT-PCR reverse transcription:
taking the purified barley cell RNA obtained in the step (4) as a template, using a Genome LabTMGeXP start-up kit of Beckman Coulter company, using a downstream primer of the designed multiplex primer as a specific primer (the sequence of the downstream primer is shown in Table 3), and taking the total RNA of the barley cell as a template to synthesize a first cDNA chain with a reaction system of 10 mu L, wherein the reaction parameters for synthesizing the first cDNA chain are set as follows: 48 ℃ for 1 minute; 60 minutes at 42 ℃; 5 minutes at 95 ℃;
then, the method is carried out by using a DNA polymerase of BeckmanCoulter company and a GenomeLabTMGeXP start kit. RT-PCR amplification was performed using the first strand of cDNA synthesized as described above as a template and the forward primer of the 13 multiplex primers as a specific primer (see Table 3 for the sequence). The RT-PCR amplification parameters were set as follows: pre-denaturation at 95 ℃ for 10min; denaturation at 94 ℃ for 30 seconds; the annealing temperature is 56 ℃, and the annealing time is 30 seconds; extending at 71 ℃ for 1 minute, circulating for 35 times, and finally obtaining a reverse transcription amplification product of the expression gene.
TABLE 3 sequences of upstream and downstream primers
(6) Analysis of gene expression level:
and (3) carrying out quantitative analysis on the reverse transcription product by utilizing capillary electrophoresis. Adding 1 μ l of PCR multiplex product into the holes of the upper sample plate filled with 39 μ l of 95% deionized formamide (SLS) and 400bp Marker mixed solution, uniformly mixing by using a pipette gun, and covering a drop of paraffin oil;
and adding 250 mul of separation buffer solution into each hole of the buffer solution plate, performing capillary electrophoresis on the plate after all the separation buffer solution is prepared, and obtaining the expression quantity of different genes at different stages after the electrophoresis is finished.
(7) PYF performance prediction:
the gene expression of sample a and the gene expression of standard barley were subjected to PCA analysis (experimental results are shown in table 4):
TABLE 4 Gene expression PCA analysis of batch A malt and Standard malt
The principal component analysis revealed that the standard malt had a score of-1.563 and malt A had a score of-1.628, the absolute difference between these two values was 0.065 and less than 0.5, and the PYF value of malt A was 99%.
Example 4
The embodiment provides a method for predicting the PYF performance of malt based on barley gene expression in the malting process, and the embodiment predicts the PYF performance of a newly purchased batch B of barley, and specifically comprises the following steps:
the specific prediction method was the same as example 3, except that the malt lot was different.
The gene expression of batch B and that of standard barley were subjected to PCA analysis (experimental results are shown in table 5):
TABLE 5 Gene expression PCA analysis of batch B malt and Standard malt
The principal component analysis shows that the standard malt has a principal component score of-1.563 and the malt B has a principal component score of 0.345, the absolute value of the difference between the two is 1.908 which is greater than 0.5 and less than 5, and the PYF value of the malt B is 62%.
Example 5
The embodiment provides a method for predicting the PYF performance of malt based on barley gene expression in the malting process, and the embodiment predicts the PYF performance of a newly purchased batch C of barley, and specifically comprises the following steps:
the specific prediction method was the same as example 3, except that the malt lot was different.
The gene expression of lot C and that of the standard barley were subjected to PCA analysis (experimental results are shown in table 6):
TABLE 6 analysis of Gene expression PCA of malt C and Standard malt
Through principal component analysis, the standard malt principal component score is-1.563, the malt C principal component score is 3.378, the absolute value of the difference between the two is 5.121 which is more than 5.0, and the PYF value of the malt C is 41%.
Proof test
In order to further verify the accuracy of the malt PYF performance prediction method based on barley gene expression in the malting process, the traditional evaluation method is adopted for result verification, and the method specifically comprises the following steps:
the results of the conventional evaluation method (i.e., a method of preparing malt into wort, then adding yeast for fermentation, and then measuring the number of suspended yeast in the fermentation broth) were compared with those of examples 3 to 5, and the results were as follows:
the traditional evaluation method specifically comprises the following steps:
(1) Preparing wort: crushing a certain amount of malt, adding water for saccharification, and filtering to obtain saccharified wort;
(2) Yeast inoculation and fermentation: placing a certain amount of yeast paste in sterilized wort, vibrating and oxygenating, and then fermenting at 20 ℃ for 48h;
(3) And (3) counting yeasts: taking a certain amount of fermentation liquor, and counting yeasts by using a yeast counter. Obtaining the yeast concentration in the fermentation liquor;
(4) Calculating the PYF value: PYF value = number of yeast cells of sample/number of yeast cells of standard sample × 100%.
TABLE 7 comparison of conventional evaluation methods with the results of examples 3-5
As can be seen from the data in Table 7, compared with the conventional evaluation method, the method provided by the invention has the advantages that the tracking genes are selected by comparing the gene expression difference of the standard malt and the PYF malt in the malting process and combining the large production process, and the accurate prediction of the PYF performance of the malt can be realized by comparing the expression quantity conditions of the standard malt and the malt to be tested and combining the evaluation standard.
Claims (10)
1. A malt PYF performance prediction method based on barley gene expression in a malting process is characterized by comprising the following steps:
analyzing the gene expression difference of the standard malt and the PYF malt in the malting process through a transcriptome, and selecting a plurality of genes with larger expression difference as candidate genes;
tracking the mass production process of the standard malt and the PYF malt, and further selecting the candidate genes with the most obvious change trend as tracking genes;
designing a reverse transcription reaction primer group and a multiple PCR reaction primer group according to the tracked gene sequence;
extracting total RNA of the malt to be detected and the standard malt;
taking the extracted total RNA as a template, and taking the reverse transcription reaction primer group as a primer to carry out reverse transcription reaction to prepare a cDNA template;
and (3) performing multiple PCR reaction by taking the reverse transcription reaction product as a template and the multiple PCR reaction primer group as a primer, and performing electrophoresis and expression analysis on the multiple PCR reaction products of the malt to be detected and the standard malt to predict the PYF performance of the malt.
2. The method of claim 1, wherein the candidate gene is selected by transcriptionally analyzing differences in gene expression between the standard malt and the PYF malt during germination during malting.
3. The method of claim 1, wherein the screening criteria for the trace genes are:
the variation amplitude of gene expression is more than or equal to 2.0 or less than 0.5;
wherein the gene expression variation amplitude is the ratio of the PYF malt gene expression principal component score to the standard malt gene expression principal component score.
4. The malt PYF performance prediction method based on malting process barley gene expression of claim 1, characterized in that, the trace genes include 13 genes including AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17 and WRKY 5.
5. The malt PYF performance prediction method based on barley gene expression in malting process of claim 4, characterized in that, the reverse transcription primer group is designed to obtain 13 primers aiming at 13 tracking genes AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17 and WRKY5, and the primers are respectively shown as SEQ ID NO.2, SEQ ID NO.4, SEQ ID NO.6, SEQ ID NO.8, SEQ ID NO.10, SEQ ID NO.12, SEQ ID NO.14, SEQ ID NO.16, SEQ ID NO.18, SEQ ID NO.20, SEQ ID NO.22, SEQ ID NO.24 and SEQ ID NO. 26.
6. The malt PYF performance prediction method based on barley gene expression in malting process of claim 4, characterized in that, the multiple PCR reaction primer set is designed to obtain 13 primers aiming at 13 tracking genes AP1, WRKY70, OSM1, GA2ox3, thioredoxin, serpin, RPP13, rbohE, rbohB2, A2, IAA16, BGU17 and WRKY5, and the primers are respectively shown as SEQ ID NO.1, SEQ ID NO.3, SEQ ID NO.5, SEQ ID NO.7, SEQ ID NO.9, SEQ ID NO.11, SEQ ID NO.13, SEQ ID NO.15, SEQ ID NO.17, SEQ ID NO.19, SEQ ID NO.21, SEQ ID NO.23 and SEQ ID NO. 25.
7. The method of claim 1, wherein the performance of PYF is determined by performing electrophoresis and expression analysis on multiple PCR reaction products of the malt to be tested and standard malt according to the following criteria:
obtaining the principal component scores of the standard malt and the malt to be detected through principal component analysis, and taking the absolute value of the difference between the two scores;
when the absolute value of the difference value of the two is less than 0.5, the PYF value of the malt to be detected is more than or equal to 90 percent;
when the absolute value of the difference value of the two is more than 0.5 and less than 5.0, the PYF value of the malt to be detected is 50-90 percent;
when the absolute value of the difference value of the two is more than 5.0, the PYF value of the malt to be detected is less than 50 percent.
8. The method of claim 7, wherein the PYF performance criteria is determined by:
performing gene expression analysis on the tracking genes of different PYF malts to obtain a gene expression principal component score;
comparing the gene expression principal component score to a gene expression principal component score of a standard malt, thereby establishing the malt PYF performance criterion.
9. Use of a malt PYF performance prediction method based on barley gene expression during malting according to any one of claims 1-8 for the prediction of malt PYF performance.
10. Use of a malt PYF performance prediction method based on malting barley gene expression in the prediction of malt PYF performance according to claim 9, wherein the malt includes finished malt, barley and malted barley malt germinated in malting.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2024037669A1 (en) * | 2022-11-24 | 2024-02-22 | 青岛啤酒股份有限公司 | Method for regulating and controlling malt pyf on basis of gene expression in malting process and use thereof |
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