CN116121092A - Recombinant saccharomyces cerevisiae with enhanced multiple stress resistance, construction method and application thereof - Google Patents
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Abstract
The invention discloses a recombinant saccharomyces cerevisiae with enhanced multiple stress resistance, and a construction method and application thereof, and belongs to the technical field of bioengineering. The invention clones the gene (OLE 1) or (FAD 2) of desaturase for controlling unsaturated fatty acid synthesis to the salt-tolerant saccharomyces cerevisiae (Zygosaccharomyces rouxii 3791), and converts the saccharomyces cerevisiae (Saccharomyces cerevisiae CEN.PK2-1C) by connecting to a bidirectional expression plasmid pY15TEF-1 capable of high copy number in the saccharomyces cerevisiae, thus obtaining the saccharomyces cerevisiae genetic engineering bacteria with improved salt tolerance, acid tolerance, heat resistance and cold resistance.
Description
Technical Field
The invention relates to the technical field of bioengineering, in particular to recombinant saccharomyces cerevisiae with enhanced multiple stress resistance, and a construction method and application thereof.
Background
Microorganisms are susceptible to a variety of environmental disturbances during fermentation and growth, including exogenous and endogenous environments. The Russell microzyme is a microzyme with high osmotic pressure resistance, can grow in materials with high sugar content and high salt content, and can not completely inhibit the growth even under saturated salt condition. The rupeste is an important strain for brewing soy sauce and sauce, and can endow the products with aroma components such as ethanol, esters, furfural, succinic acid, furanone and the like. Therefore, it is widely used in the production process of conventional high-salt fermented foods such as bean paste, soy sauce, miso, etc. Saccharomyces cerevisiae (Saccharomyces cerevisiae) is an important industrial model strain, however, adverse environmental factors often make it difficult for Saccharomyces cerevisiae to maintain high strength fermentative production.
In view of this, the present invention has been made.
Disclosure of Invention
The invention aims to provide a recombinant saccharomyces cerevisiae with enhanced multiple stress resistance, a construction method and application thereof, and the unsaturated fatty acid synthesis capability can be improved by over-expressing a desaturase gene for controlling unsaturated fatty acid synthesis in the saccharomyces cerevisiae, so that the multiple stress resistance of the saccharomyces cerevisiae is enhanced.
In order to achieve the aim, the invention constructs the recombinant saccharomyces cerevisiae with enhanced multiple stress resistance. The invention firstly introduces the vector containing desaturase genes of the roux yeast into the saccharomyces cerevisiae, and discovers that the unsaturated fat synthesis capability of recombinant bacteria introduced with the vector containing the desaturase genes is improved, thereby enhancing the multiple stress tolerance capability of the recombinant bacteria.
Specifically, the invention provides the following technical scheme:
in a first aspect, the present invention provides a recombinant saccharomyces cerevisiae with enhanced resistance to multiple stresses, the recombinant saccharomyces cerevisiae comprising exogenous genes;
the exogenous genes include at least one desaturase gene that controls unsaturated fatty acid synthesis;
such multiple stress resistance includes resistance to salt stress, acid stress, heat stress and cold stress.
The invention discovers that the recombinant saccharomyces cerevisiae containing the desaturase gene can enhance the resistance in the aspects of salt stress, acid stress, heat stress, cold stress and the like, so that the constructed recombinant saccharomyces cerevisiae can overcome adverse environmental factors in fermentation production.
In some embodiments, the desaturase gene is derived from saccharomyces rouxii.
In some embodiments, the foregoing rouxii is rouxii with a preservation number of CGMCC No.3791, a preservation time of 4 months and 29 days in 2010, a preservation place of the general microbiological center of the China Committee for culture Collection of microorganisms, and an address of the general microbiological center of the Beijing area, the Chaoyang area, the Beichen, no.1, and No. 3.
In some embodiments, desaturase genes include OLE1 genes and FAD2 genes.
In some embodiments, the NCBI gene IDs of the OLE1 gene and FAD2 gene are zygr_0ai02130 and zygr_0ag03090, respectively.
In some embodiments, the nucleotide sequences of the OLE1 gene and the FAD2 gene are SEQ ID nos. 1-2.
In some embodiments, the starting strain of recombinant Saccharomyces cerevisiae is Saccharomyces cerevisiae CEN.PK2-1C, disclosed in van Di jken JP et al, enzyme microb.technology.2000, 26:706-714, genotype MATA ura3-52leu3-5,112trp1-289 his3ΔMAL2-8 cSUC2, a benefit of university of Jiangnan.
In some embodiments, the recombinant saccharomyces cerevisiae described above contains a high copy bidirectional expression plasmid.
In some embodiments, the expression plasmid described above comprises pY15TEF-1 (Biovector co., LTD) with a leucine LEU tag, offered by university of south of the river.
In a second aspect, the present invention provides a method for constructing the recombinant saccharomyces cerevisiae, comprising: and connecting the desaturase gene to an expression plasmid, and then introducing the recombinant expression plasmid into saccharomyces cerevisiae to obtain the recombinant saccharomyces cerevisiae.
Specifically, the construction method comprises the following steps:
(1) The target gene OLE1 with two restriction sites of HindIII and XhoI and a protective base at two ends and the target gene FAD2 with two restriction sites of BamH I and XhoI and a protective base at two ends are respectively connected to pY15TEF1 which is subjected to the same restriction enzyme, and E.coli DH5a is respectively transformed, and the obtained transformant is identified as recombinant plasmids pY15TEF1-OLE1 and pY15TEF1-FAD2 by colony PCR verification, extraction plasmid restriction enzyme verification and sequencing.
(2) And (3) transforming the constructed recombinant plasmid into Saccharomyces cerevisiae CEN.PK2-1C to obtain the recombinant Saccharomyces cerevisiae with improved multiple stress resistance.
In a third aspect, the invention provides the use of the recombinant Saccharomyces cerevisiae described above in the field of fermentation brewing and fermented food.
In a fourth aspect, the present invention provides a method for enhancing multiple stress resistance of Saccharomyces cerevisiae comprising using the recombinant Saccharomyces cerevisiae to overexpress a desaturase gene to control unsaturated fatty acid synthesis.
In some embodiments, the overexpression is performed by constructing a recombinant plasmid containing a gene encoding a desaturase through the gene encoding the desaturase and an expression plasmid, and then introducing the recombinant plasmid into Saccharomyces cerevisiae to induce expression of the desaturase.
In some embodiments, the multiple stress resistance comprises resistance to salt stress, acid stress, heat stress, and cold stress.
In some embodiments, the Saccharomyces cerevisiae is Saccharomyces cerevisiae CEN.PK2-1C, which has a genotype of MATA ura3-52leu3-5,112trp1-289 his3ΔMAL2-8C SUC2.
In some embodiments, the desaturase gene is derived from saccharomyces rouxii.
In some embodiments, the collection number of the saccharomyces rouxii is CGMCC No.3791.
In some embodiments, desaturase genes include OLE1 genes and FAD2 genes.
In some embodiments, the NCBI gene IDs of the OLE1 gene and FAD2 gene are zygr_0ai02130 and zygr_0ag03090, respectively.
In some embodiments, the nucleotide sequences of the OLE1 gene and the FAD2 gene are SEQ ID nos. 1-2.
In some embodiments, the expression plasmid is a high copy bidirectional expression plasmid.
In some embodiments, the expression plasmid comprises pY15TEF-1.
The invention has the following beneficial effects:
according to the invention, the expression of the OLE1 or FAD2 genes of the saccharomyces cerevisiae is cloned and introduced into the saccharomyces cerevisiae, so that the expression of desaturase genes is induced, and the unsaturation degree and multiple stress tolerance of the fatty acid of the saccharomyces cerevisiae are enhanced.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings that are needed in the embodiments will be briefly described below, it being understood that the following drawings only illustrate some embodiments of the present invention and therefore should not be considered as limiting the scope, and other related drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 shows the results of the fatty acid unsaturation in examples 1-2 and comparative example 1.
FIG. 2 shows the statistics of viable cell count under salt stress conditions of examples 1-2 and comparative example 1.
FIG. 3 shows the statistics of viable cell count under acid stress conditions of examples 1-2 and comparative example 1.
FIG. 4 shows the statistics of viable count under heat stress conditions of examples 1-2 and comparative example 1.
FIG. 5 shows the statistics of viable cell count under cold stress conditions of examples 1-2 and comparative example 1.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention more clear, the technical solutions of the embodiments of the present invention will be clearly and completely described below. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
Example 1
The embodiment provides a recombinant saccharomyces cerevisiae and a construction method thereof.
Wherein, the exogenous genes in the recombinant saccharomyces cerevisiae are: the OLE1 gene with the nucleotide sequence of SEQ ID NO. 1. The host is: saccharomyces cerevisiae CEN.PK2-1C; the vector is pY15TEF-1, and is a benefit of Jiang Nada.
The construction method comprises the following steps:
(1) Extracting total RNA of the Rumex rouxii (Z. Rouxii CGMCC No. 3791), and obtaining a desaturase gene OLE1 with two enzymatic cleavage sites of Xho I and Hind III at two ends by PCR. The primers used for cloning OLE1 were:
F1:CCCTCGAGATGAGTTCAATGGAGGAGGTGGAC，SEQ ID NO.3；
R1:CCCAAGCTTTTACTGAGCCTTCTTGGCAGAGTAG，SEQ ID NO.4。
(2) Then, the recombinant plasmid pY15TEF1-OLE1 is obtained by connecting the recombinant plasmid pY15TEF1 plasmid to the plasmid subjected to the same digestion, transforming the escherichia coli DH5 alpha, obtaining the transformant, and performing colony PCR verification, plasmid digestion verification extraction and sequencing verification.
(3) And transforming Saccharomyces cerevisiae (S.cerevisiae CEN.PK2-1C) with the constructed recombinant plasmid, extracting plasmids of the transformants, and performing enzyme digestion verification to confirm positive transformants.
Example 2
The embodiment provides a recombinant saccharomyces cerevisiae and a construction method thereof.
Wherein, the exogenous genes in the recombinant saccharomyces cerevisiae are: FAD2 gene with nucleotide sequence of SEQ ID NO. 2. The host was the same as the vector in example 1.
The construction method comprises the following steps:
(1) Extracting total RNA of the Rumex rouxii (Z.rouxii CGMCC No. 3791), and obtaining a desaturase gene FAD2 with two enzyme cutting sites of BamH I and Xho I by PCR, wherein primers used for cloning the FAD2 are as follows:
F2:CCCTCGAGATGACCGTTAACAGGACGACCAGT，SEQ ID NO.5；
R2:CGGGATCCTTATGACCTCTTCTCCTTCTCTGCGC，SEQ ID NO.6。
(2) Then connecting to pY15TEF1 plasmid subjected to same digestion, transforming escherichia coli DH5 alpha, obtaining transformant, and carrying out colony PCR verification, plasmid digestion verification and sequencing to confirm the recombinant plasmid pY15TEF1-FAD2.
(3) And transforming Saccharomyces cerevisiae (S.cerevisiae CEN.PK2-1C) with the constructed recombinant plasmid, extracting plasmids of the transformants, and performing enzyme digestion verification to confirm positive transformants.
Comparative example
The recombinant Saccharomyces cerevisiae constructed in this comparative example was: the empty plasmid pY15TEF1 was transformed into Saccharomyces cerevisiae (S.cerevisiae CEN.PK2-1C), and the plasmid of the transformant was extracted and verified by cleavage, and confirmed as a positive transformant.
Experimental example 1
The experimental example is for measuring the content of unsaturated fatty acid, and comprises the following specific steps:
(1) The strain of examples 1-2 and comparative example, which was stored in glycerol stock at-80 ℃, was taken: recombinant Saccharomyces cerevisiae S.PK2-1C pY15TEF1-OLE1, S.cerevisiae CEN.PK2-1C pY15TEF1-FAD2 and S.cerevisiae CEN.PK2-1CpY15TEF1 were inoculated in seed medium at an inoculum size of 10% (V/V) and cultured at 30℃for 12 hours to mid-log phase.
The seed culture medium comprises the following components: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L.
(2) The cells were collected for unsaturated fatty acid content determination and methods for unsaturated fatty acid extraction and determination were published in Wu et al, j.ind.microbiol.biotechnol.2012, 39:1031-1039. The results are shown in FIG. 1.
From fig. 1, it can be seen that after overexpression of OLE1 or FAD2, the fatty acid unsaturation was increased by 2.6% and 11.7%, respectively, compared to the starting strain.
Experimental example 2
The experimental example is the detection of the resistance under the condition of salt stress, and comprises the following specific steps:
(1) The strain of examples 1-2 and comparative example, which was stored in glycerol stock at-80 ℃, was taken: recombinant Saccharomyces cerevisiae S.PK2-1C pY15TEF1-OLE1, S.cerevisiae CEN.PK2-1C pY15TEF1-FAD2 and S.cerevisiae CEN.PK2-1CpY15TEF1 were inoculated into a seed culture medium in an inoculum size of 10% (V/V), and were subjected to stationary culture at 30℃for 12 hours to mid-log phase to obtain a seed culture solution.
The seed culture medium comprises the following components: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L.
(2) Seed culture broth was inoculated into salt stress medium at an inoculum size of 10% (V/V) for 4h.
The components of the salt stress culture medium comprise: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L, sodium chloride NaCl 70.2g/L.
(3) Centrifuging the culture solution subjected to salt stress at 8000r/min (4deg.C) for 5min, collecting thallus, suspending the thallus in sterile water, and controlling initial biomass to 0.5OD 600 After dilution by 1000 times, 5. Mu.L of the culture medium was applied to a solid plate medium, and the culture was allowed to stand at 30℃for 48 hours to calculate the colony count, and the result was shown in FIG. 2.
The solid medium comprises the following components: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L, agar 20g/L.
As can be seen from FIG. 2, after over-expression of OLE1 or FAD2, the viable count of OLE1 or FAD2 strain was increased by 13% and 30.4%, respectively, compared to the starting strain. From this, it was demonstrated that the unsaturated fatty acid synthesis ability can be improved by overexpressing a desaturase gene controlling unsaturated fatty acid synthesis in Saccharomyces cerevisiae, thereby enhancing the salt stress tolerance of recombinant bacteria.
Experimental example 3
The experimental example is the detection of the resistance under the condition of acid stress, and comprises the following specific steps:
(1) The strain of examples 1-2 and comparative example, which was stored in glycerol stock at-80 ℃, was taken: recombinant Saccharomyces cerevisiae S.PK2-1C pY15TEF1-OLE1, S.cerevisiae CEN.PK2-1C pY15TEF1-FAD2 and S.cerevisiae CEN.PK2-1C pY15TEF1 are inoculated into a seed culture medium in an inoculum size of 10% (V/V), and the seed culture medium is obtained by standing culture at 30 ℃ for 12 hours to mid-log phase.
The seed culture medium comprises the following components: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L.
(2) The seed culture was inoculated into an acid stress medium at an inoculum size of 10% (V/V) for 4 hours.
The acid stress medium comprises the following components: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L, acetic acid 5g/L.
(3) Centrifuging the culture solution subjected to acid stress at 8000r/min (4deg.C) for 5min, collecting thallus, suspending the thallus in sterile water, and controlling initial biomass to 0.5OD 600 After dilution by 1000 times, 5. Mu.L of the culture medium was applied to a solid plate medium, and the culture was allowed to stand at 30℃for 48 hours to calculate the colony count, and the result was shown in FIG. 3.
The solid medium comprises the following components: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L, agar 20g/L.
As can be seen from FIG. 3, after over-expression of OLE1 or FAD2, the viable count of OLE1 or FAD2 strain was increased by 26.1% and 39.1%, respectively, compared to the starting strain. From this, it was demonstrated that the unsaturated fatty acid synthesis ability can be improved by overexpressing a desaturase gene controlling unsaturated fatty acid synthesis in Saccharomyces cerevisiae, thereby enhancing the acid stress tolerance of recombinant bacteria.
Experimental example 4
The experimental example is the detection of the resistance under the heat stress condition, and comprises the following specific steps:
(1) The strain of examples 1-2 and comparative example, which was stored in glycerol stock at-80 ℃, was taken: recombinant Saccharomyces cerevisiae S.PK2-1C pY15TEF1-OLE1, S.cerevisiae CEN.PK2-1C pY15TEF1-FAD2 and S.cerevisiae CEN.PK2-1CpY15TEF1 were inoculated into a seed culture medium in an inoculum size of 10% (V/V), and were subjected to stationary culture at 30℃for 12 hours to mid-log phase to obtain a seed culture solution.
(2) The seed culture was inoculated into the seed medium at an inoculum size of 10% (V/V), and treated at 40℃for 4 hours.
The seed culture medium comprises the following components: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L.
(3) Centrifuging the culture solution subjected to heat stress treatment (40 ℃ C. For 4 hr) at 8000r/min (4 ℃) for 5min, collecting thallus, suspending the thallus in sterile water, and controlling initial biomass to 0.5OD 600 After dilution by 1000 times, 5. Mu.L of the culture medium was applied to a solid plate medium, and the culture was allowed to stand at 30℃for 48 hours to calculate the colony count, and the result is shown in FIG. 4.
The solid medium comprises the following components: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L, agar 20g/L.
As can be seen from FIG. 4, after over-expression of OLE1 or FAD2, the viable count of OLE1 or FAD2 strain was increased by 7.1% and 17.9%, respectively, compared to the starting strain. From this, it was demonstrated that the unsaturated fatty acid synthesis ability can be improved by overexpressing a desaturase gene controlling unsaturated fatty acid synthesis in Saccharomyces cerevisiae, thereby enhancing the heat stress tolerance of recombinant bacteria.
Experimental example 5
The experimental example is the detection of the resistance under the condition of cold stress, and comprises the following specific steps:
(1) The strain of examples 1-2 and comparative example, which was stored in glycerol stock at-80 ℃, was taken: recombinant Saccharomyces cerevisiae S.PK2-1C pY15TEF1-OLE1, S.cerevisiae CEN.PK2-1C pY15TEF1-FAD2 and S.cerevisiae CEN.PK2-1CpY15TEF1 were inoculated into a seed culture medium in an inoculum size of 10% (V/V), and were subjected to stationary culture at 30℃for 12 hours to mid-log phase to obtain a seed culture solution.
(2) Seed culture broth was inoculated into the seed medium at an inoculum size of 10% (V/V), and treated at 4℃for 4 hours.
The seed culture medium comprises the following components: 6.7g/L yeast nitrogen source free medium (YNB does not contain (NH 4) 2SO 4), 20g/L glucose, 0.02g/L tryptophan Try, 0.02g/L histidine His and 0.02g/L uracil Ura.
(3) Centrifuging the culture solution subjected to cold stress treatment (4 deg.C for 4 hr) at 8000r/min (4 deg.C) for 5min, collecting thallus, suspending the thallus in sterile water, and controlling initial biomass to 0.5OD 600 After dilution by 1000 times, 5. Mu.L of the culture medium was applied to a solid plate medium, and the culture was allowed to stand at 30℃for 48 hours to calculate the colony count, and the result was shown in FIG. 5.
The solid medium comprises the following components: 6.7g/L yeast-free nitrogen source medium (YNB does not contain (NH) 4 ) 2 SO 4 ) Glucose 20g/L, tryptophan Try 0.02g/L, histidine His0.02g/L, uracil Ura 0.02g/L, agar 20g/L.
As can be seen from FIG. 5, after over-expression of OLE1 or FAD2, the viable count of OLE1 or FAD2 strain was increased by 47.6% and 33.3%, respectively, compared to the starting strain. From this, it was demonstrated that the unsaturated fatty acid synthesis ability can be improved by overexpressing a desaturase gene controlling unsaturated fatty acid synthesis in Saccharomyces cerevisiae, thereby enhancing the cold stress tolerance of recombinant bacteria.
The above description is only of the preferred embodiments of the present invention and is not intended to limit the present invention, but various modifications and variations can be made to the present invention by those skilled in the art. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (10)
1. The recombinant saccharomyces cerevisiae with enhanced multiple stress resistance is characterized in that the recombinant saccharomyces cerevisiae contains exogenous genes; the exogenous genes include at least one desaturase gene that controls unsaturated fatty acid synthesis;
the multiple stress resistance includes resistance to salt stress, acid stress, heat stress and cold stress.
2. The recombinant saccharomyces cerevisiae according to claim 1, wherein said desaturase gene is derived from saccharomyces rouxii;
preferably, the preservation number of the rupeste is CGMCC No.3791.
3. The recombinant saccharomyces cerevisiae according to claim 2 wherein said desaturase genes include OLE1 and FAD2 genes;
preferably, the NCBI gene IDs of the OLE1 gene and the FAD2 gene are ZYGR_0AI02130 and ZYGR_0AG03090, respectively;
preferably, the nucleotide sequences of the OLE1 gene and the FAD2 gene are SEQ ID NO.1-2.
4. A recombinant saccharomyces cerevisiae according to claim 3 wherein the starting strain of the recombinant saccharomyces cerevisiae is saccharomyces cerevisiae cen.pk2-1C with genotype MATa ura3-52leu3-5,112trp1-289 his3Δmal 2-8C sud 2.
5. The recombinant saccharomyces cerevisiae according to claim 4 wherein said recombinant saccharomyces cerevisiae contains high copy bidirectional expression plasmids;
preferably, the expression plasmid comprises pY15TEF-1.
6. The method for constructing a recombinant saccharomyces cerevisiae according to any of the claims 1-5, wherein said method for constructing comprises: and connecting the desaturase gene to the expression plasmid, and then introducing the recombinant expression plasmid into saccharomyces cerevisiae to obtain the recombinant saccharomyces cerevisiae.
7. Use of the recombinant saccharomyces cerevisiae according to any of claims 1-5 in the field of fermentation brewing and fermented food.
8. A method of enhancing multiple stress resistance of saccharomyces cerevisiae, comprising over-expressing a desaturase gene using the recombinant saccharomyces cerevisiae of any of claims 1-5 to control unsaturated fatty acid synthesis;
preferably, the over-expression is that a gene recombinant plasmid containing the coding desaturase is constructed through the gene coding the desaturase and an expression plasmid, and then the recombinant plasmid is introduced into saccharomyces cerevisiae to induce and express the desaturase;
preferably, the multiple stress resistance comprises resistance to salt stress, acid stress, heat stress and cold stress.
9. The method for enhancing multiple stress resistance of saccharomyces cerevisiae according to claim 8, wherein the saccharomyces cerevisiae is saccharomyces cerevisiae cen.pk2-1C with genotype MATa ura3-52leu3-5,112trp1-289 his3Δmal2-8C sud 2.
10. The method of enhancing multiple stress resistance of saccharomyces cerevisiae according to claim 9, wherein the desaturase gene is derived from saccharomyces rouxii;
preferably, the preservation number of the rupeste is CGMCC No.3791;
preferably, the desaturase genes include OLE1 genes and FAD2 genes;
preferably, the NCBI gene IDs of the OLE1 gene and the FAD2 gene are ZYGR_0AI02130 and ZYGR_0AG03090, respectively;
preferably, the nucleotide sequences of the OLE1 gene and the FAD2 gene are SEQ ID NO.1-2;
preferably, the expression plasmid is a high copy bidirectional expression plasmid;
preferably, the expression plasmid comprises pY15TEF-1.
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