A Gene Recombinant Saccharomyces cerevisiae with High Yield of Sugarcane Molasses Alcohol
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A technology of sugar cane molasses and Saccharomyces cerevisiae, applied in the fields of biotechnology and bioengineering
Active Publication Date: 2020-08-11
GUANGXI ACAD OF SCI
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Problems solved by technology
At present, industrial production is still a technical problem for the production of alcohol by fermentation of non-fermentable sugar from sugarcane molasses
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Embodiment 1
[0009] Embodiment 1: the construction of genetic engineering strain
[0010] α-galactosidase gene agl3 of Trichoderma reesei was amplified by PCR and connected to the vector pYX212 to obtain pYX212-agl3;
[0011] Primers were designed according to the TPI promoter and terminator sequences located on both sides of the reading frame on pYX212, and the yeast genome homologous integration sequence CCACAGCAGT was added to the 5' end of the primer, and the TPI promoter was amplified using pYX212-agl3 as a template The target gene TPIP-agl3-TPIT of subson and terminator;
[0012] The TPIP-agl3-TPIT fragment was electrotransformed into the haploid cells of wild-type Saccharomyces cerevisiae, and the transformed bacterial solution was inserted into the double medium for overnight culture, and the YPD plate was coated, and a single colony was inserted into the liquid YPR (1% Yeast In Extract, 2% Peptone, 2% Raffinose) medium, use a microplate reader to measure the absorbance value of t...
Embodiment 2
[0015] Embodiment 2: The growth rate of the high-yield bacterial strain that screens out in the sugarcane molasses culture medium of different brix, alcohol fermentation concentration, the mensuration of sugar utilization ability
[0016] The raw material sugarcane molasses used in the present invention conforms to the QB / T2685-2005 standard, and YPD medium is first used during fermentation (dissolve 20g Peptone, 10g Yeast Extract in 900mL water, autoclave at 121°C for 20min, add 100mL20gGlucose after sterilization alone) Activation of bacterial strains is inserted into the molasses fermentation liquid that concentration is respectively 10Bx °, 20Bx °, 30Bx °, 40Bx ° by the inoculum size of 5% (V / V) (the molasses is diluted to a certain degree of brix, add 0.2% (W / W) urea and 0.02% (W / W) phosphoric acid, with concentrated H 2 SO 4 Adjust the pH value to 3.8-4.0, ready to use), 30 ° C, 180 rpm culture, and measure the number of bacteria, ethanol concentration, total sugar con...
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Abstract
The invention relates to a genetic recombination brewer's yeast TGS11 for achieving high yield of cane molasses alcohol. The strain is preserved in the China Center for Type Culture Collection at minus 70 DEG C with the preservation number being CCTCC NO:M2013562, and the preservation date is Nov 10th, 2013. The brewer's yeast strain is characterized in that an alpha-galactosidase gene of trichoderma reesei is introduced into a wild type brewer's yeast, the gene is recombined with a genome of the strain after being introduced, and the brewer's yeast strain is obtained through screening; the yield of alcohol obtained through fermentation of the strain TGS11 in a culture medium containing 27% of saccharose and 3% of raffinose can reach 16.18% maximumly, and the yield of alcohol reaches 14% when 30Bx-degree molasses and 50Bx-degree molasses are mixed for fermentation at 30 DEG C under 180-200 rpm. The brewer's yeast strain has the advantages that an exogenous gene is successfully introduced into the wild type brewer's yeast, high expression is achieved, the strain can efficiently ferment cane molasses to produce alcohol, and good alcohol fermentation production potential is achieved.
Description
technical field [0001] The invention belongs to the field of biotechnology and bioengineering, and specifically relates to the genetic modification of Saccharomyces cerevisiae and the selection and breeding of sugarcane molasses alcoholic fermentation strains. Background technique [0002] Biomass resources are recognized as the largest renewable resources on the earth. With the depletion of non-renewable resources such as petroleum, the research on renewable biomass resources has become more and more extensive and in-depth, and has risen to the national sustainable development strategy high. Cane molasses is a by-product of the sucrose production process, which contains about 50% sugar, and is the main raw material for alcohol fermentation in sucrose producing areas such as Guangxi, southern Guangdong and Yunnan. The production of alcohol by fermentation of sugarcane molasses can not only solve the problem of environmental pollution, but also reduce the consumption of grai...
Claims
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