Genetically engineered bacterium co-generating geraniol and nerol and construction method and application thereof

A technology of genetically engineered bacteria and nerol, applied in the field of genetic engineering, can solve problems affecting the quality of geraniol and downstream applications, environmental pollution by chemical methods, and many impurities in products, and achieve easy engineering operations, cheap raw materials, and high fermentation short cycle effect

Active Publication Date: 2014-07-02
QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the chemical method has problems such as environmental pollution and harsh conditions, and the synthesized product has many impurities, which affects the quality and downstream applications of geraniol

Method used

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  • Genetically engineered bacterium co-generating geraniol and nerol and construction method and application thereof
  • Genetically engineered bacterium co-generating geraniol and nerol and construction method and application thereof
  • Genetically engineered bacterium co-generating geraniol and nerol and construction method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Example 1 Expression of phosphatidic acid phosphatase DPP1 from Saccharomyces cerevisiae for biosynthesis of nerol and geraniol

[0052] By co-expressing the acetyl-CoA acyltransferase gene / hydroxymethylglutaryl-CoA reductase gene (mvaE) from Enterococcus faecalis in Escherichia coli, 3-hydroxy-3-methylglutaryl-CoA synthesis Enzyme gene (mvaS); mevalonate kinase gene (ERG12), mevalonate-5-phosphate kinase gene (ERG8), mevalonate-5-diphosphate decarboxylase gene (ERG19) from Saccharomyces cerevisiae , isopentenyl pyrophosphate isomerase gene (IDI1); geranyl diphosphate synthase gene (GPPS2) from North American fir and phosphatidic acid phosphatase DPP1 from Saccharomyces cerevisiae (NCBI Reference Sequence: NM_001180592.1) , the biosynthesis of geraniol from glucose.

[0053] (1) Cloning of DPP1 gene and construction of expression vector

[0054] Using Saccharomyces cerevisiae S288c genomic DNA as template, DPP1-rbs-F2 and DPP1-XhoR as primers for PCR amplification, PC...

Embodiment 2

[0061] Example 2 Expression of phosphatidic acid phosphatase LPP1 from Saccharomyces cerevisiae for biosynthesis of nerol and geraniol

[0062] By co-expressing the acetyl-CoA acyltransferase gene / hydroxymethylglutaryl-CoA reductase gene (mvaE) from Enterococcus faecalis in Escherichia coli, 3-hydroxy-3-methylglutaryl-CoA synthesis Enzyme gene (mvaS); mevalonate kinase gene (ERG12), mevalonate-5-phosphate kinase gene (ERG8), mevalonate-5-diphosphate decarboxylase gene (ERG19) from Saccharomyces cerevisiae , isopentenyl pyrophosphate isomerase gene (IDI1); geranyl diphosphate synthase gene (GPPS2) from North American fir and phosphatidic acid phosphatase LPP1 from Saccharomyces cerevisiae (NCBI Reference Sequence: NM_001180811.3) , the biosynthesis of geraniol from glucose.

[0063] (1) Cloning of LPP1 gene and construction of expression vector

[0064] Using Saccharomyces cerevisiae S288c genomic DNA as template, LPP1-rbs-F2 and LPP1-XhoR as primers for PCR amplification, PC...

Embodiment 3

[0072] Example 3 Expression of ADP-ribose pyrophosphatase gene EcNudF derived from Escherichia coli for biosynthesis of nerol and geraniol

[0073] By co-expressing the acetyl-CoA acyltransferase gene / hydroxymethylglutaryl-CoA reductase gene (mvaE) from Enterococcus faecalis in Escherichia coli, 3-hydroxy-3-methylglutaryl-CoA synthesis Enzyme gene (mvaS); mevalonate kinase gene (ERG12), mevalonate-5-phosphate kinase gene (ERG8), mevalonate-5-diphosphate decarboxylase gene (ERG19) from Saccharomyces cerevisiae , isopentenyl pyrophosphate isomerase gene (IDI1); geranyl diphosphate synthase gene (GPPS2) from North American fir and ADP-ribose pyrophosphatase gene EcNudF from Escherichia coli (GenBank: U22009.1 ), using glucose to biosynthesize geraniol.

[0074] (1) Cloning of EcNudF gene and construction of expression vector

[0075] Using Escherichia coli K12 genomic DNA as a template, eNudF-rbs-BglF and eNudF-XhoR as primers for PCR amplification, PCR amplification conditions...

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Abstract

The invention discloses a genetically engineered bacterium co-generating geraniol and nerol and a construction method and an application thereof, and belongs to the technical field of generic field. According to the genetically engineered bacterium disclosed by the invention, acetyl coenzyme A acyltransferase / hydroxymethyl glutaryl coenzyme A reductase, 3-hydroxyl-3-methyl glutaryl coenzyme A synthase, mevalonic acid kinase, mevalonic acid-5-phosphokinase, mevalonic acid-5-diphosphonic acid decarboxylase, isopentenylpyrophosphate isomerase, geranyl diphosphonic acid synthetase and geraniol synthetase or phosphatase are expressed. The metabolic pathways of geraniol and nerol synthesized in escherichia coli are successfully constructed by genetic engineering means, and glucose is biologically converted into geraniol and nerol.

Description

technical field [0001] The invention relates to a genetically engineered bacterium for co-producing geraniol and nerol, its construction method and application, and belongs to the technical field of genetic engineering. Background technique [0002] Geraniol (trans-3,7-methyl-2,6-octadien-1-ol, Geraniol, also known as geraniol) and its isomer nerol (cis-3,7 -Dimethyl-2,6-octadienol, nerol) is an acyclic monoterpene alcohol compound, which is one of the main components of essential oils such as rose oil, martin balm and citronella oil. Geraniol, nerol and their esters can be used in flavors and food flavors, and are the main ingredients of rose flavors, and can also be widely used in medicine, tobacco, and food ingredients. In addition, it can also be used as a natural low-toxic insect repellant, a new type of chemical anti-cancer agent. At present, the global annual demand for nerol is about 5,000 tons, and the demand in my country is no less than 500 tons, while the world...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/21C12N15/70C12N15/74C12P7/04C12R1/125C12R1/19C12R1/22
Inventor 咸漠刘炜田宁蒋昱东
Owner QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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