High-yield 3-Methionol saccharomyces cerevisiae engineering bacteria and preparation method and application thereof
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A technology of methylthiopropanol and genetically engineered bacteria, applied in the directions of genetic engineering, microorganism-based methods, botanical equipment and methods, etc., can solve problems such as expensive natural spices, achieve conversion rate and yield improvement, and obvious technical effects Effect
Inactive Publication Date: 2013-11-27
BEIJING TECHNOLOGY AND BUSINESS UNIVERSITY
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However, due to resource and cost constraints, natural flavors are more expensive
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[0016] The present invention is a Saccharomyces cerevisiae genetically engineered bacteria with high yield of 3-methylthiopropanol and its preparation method and application. The specific implementation method (technical scheme):
[0017] (1) The ARO9 and ARO10 genes were amplified from Saccharomyces cerevisiae DNA by PCR technology, connected to the shuttle plasmid expression vector, and then the recombinant vectors were introduced into S. cerevisiae S288c to overexpress the ARO9 and ARO10 genes respectively, and the gene transcription activity was detected , analysis of transaminase, decarboxylase expression levels, metabolic flux.
[0018] (2) The ARO9 and ARO10 genes were connected to the shuttle plasmid expression vector at the same time, and then the recombinant plasmid was introduced into S. cerevisiae S288c, and the positive transformants were screened, so that the ARO9 and ARO10 genes were coupled and overexpressed.
[0019] (3) Cloning the CY3 gene in S. cerevisiae S...
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Abstract
The invention relates to high-yield 3-Methionol saccharomyces cerevisiae engineering bacteria and a preparation method and application thereof, and belongs to the technical field of biological preparation of food flavors. According to the high-yield 3-Methionol saccharomyces cerevisiae engineering bacteria and the preparation method and application thereof, the engineering bacteria is designed and constructed through the coupling excessive expression of an aminotransferase gene (ARO9) and a decarboxylase gene (ARO10) and the knock-out of a methylthio lyase gene (CY3) in the anabolic pathway of Methionol by taking Saccharomyces cerevisiae S288c as an original strain; the expression of two key enzyme genes in a main synthetic pathway is enhanced, and the expression of the CY3 gene in a side reaction pathway is reduced or eliminated; the bred engineering bacteria (converter) is subjected to multiple-time passage and stable in inheritance, so that the metabolic network and metabolic pathway of the biological synthesis of the Methionol are optimized, and the metabolic flux and yield of the Methionol are obviously increased; a Methionol flavor prepared by fermenting and converting the engineering bacteria is single in structure and chirality and high in fragrance quality.
Description
technical field [0001] The invention relates to a Saccharomyces cerevisiae genetically engineered bacterium with high yield of 3-methylthiopropanol and its preparation method and application, which belongs to the field of bio-manufacturing of edible spices, and the prepared natural 3-methylthiopropanol spices can be used for various edible flavors Deployment, widely used in food, medicine, daily chemical and other industries. Background technique [0002] 3-Methylthiopropanol (Methionol, also known as pineapple alcohol) is an important edible spice, naturally occurring in wine, beer, rice wine, whiskey, brandy, soy sauce, soybean paste, cheese, bacon, rye bread, etc. Fermented food is an important flavor substance of sesame-flavored liquor, red wine, and soy sauce, and also exists in tomatoes, potatoes, and seafood. The aroma threshold of 3-methylthiopropanol is low, and the recommended dosage in food is 0.1-10mg / kg. Its flavors are malt, sesame oil and cooked potatoes in s...
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