Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Gene CsRHMb capable of catalyzing biosynthesis of UDP-rhamnose as well as encoded protein of gene CsRHMb and applications of encoded protein

A biosynthesis and rhamnose technology, applied in the fields of molecular biology and metabolic engineering, can solve the problems of difficult purification, environmental pollution, low efficiency, etc.

Active Publication Date: 2017-07-04
ANHUI AGRICULTURAL UNIVERSITY
View PDF1 Cites 7 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The content of UDP-rhamnose in plants is extremely low, its purification is difficult, and the efficiency is low, which limits the acquisition of this compound
There are also many problems in the use of modern chemical synthesis methods, such as high synthesis costs, cumbersome processes, and the production of chemical substances that pollute the environment
Therefore, it is difficult to purify and synthesize UDP-rhamnose by traditional methods
So far, there is no commercial UDP-rhamnose on the market

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Gene CsRHMb capable of catalyzing biosynthesis of UDP-rhamnose as well as encoded protein of gene CsRHMb and applications of encoded protein
  • Gene CsRHMb capable of catalyzing biosynthesis of UDP-rhamnose as well as encoded protein of gene CsRHMb and applications of encoded protein
  • Gene CsRHMb capable of catalyzing biosynthesis of UDP-rhamnose as well as encoded protein of gene CsRHMb and applications of encoded protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1 Cloning and expression of the gene CsRHMb that efficiently catalyzes UDP-rhamnose biosynthesis

[0030] 1. Materials

[0031] 1. Tea tree species: Nongkangzao (Camellia sinensis (L.) O.Kuntze.var.sinensis cultivar Nongkangzao), fresh tea leaves were collected, quickly frozen with liquid nitrogen, and stored in a -80°C refrigerator for later use;

[0032] 2. Escherichia coli Novablue (DE3) expression host bacteria: purchased from Shanghai Beinuo Biotechnology Co., Ltd.;

[0033] 3. LB medium: Weigh 10g NaCl, 5g yeast extract, 10g tryptone, add 950mL ultrapure water and stir to dissolve, adjust pH to 7.0 with 1mol / L NaOH, add water to 1000mL, autoclave for 15min, Promptly obtain LB liquid medium, LB solid medium is to add 15g agar powder in LB liquid medium;

[0034] 4. Glucose solution with a mass ratio of 40%: Weigh 40g of glucose, add ultrapure water to dissolve and stir evenly, set the volume to 100mL, and sterilize at 110°C for 10min;

[0035] 5. Ampicil...

Embodiment 2

[0054] Example 2 Detection and Analysis of Enzymatic Activity of rCsRHMb Recombinant Protein

[0055] 1. Enzyme activity detection of UDP-glucose substrate

[0056] The total reaction volume is 50 μL, 100 mM NaH 2 PO 4 / Na 2 HPO 4 (pH: 9.5) buffer solution contains 3mM UDP-glucose as reaction substrate, 3mM NAD and 3mM NASPH as coenzyme, 10-20μg purified recombinant protein rCsRHMb.

[0057] After the above enzyme reaction system was bathed in water at 35°C for 1 hour, the reaction was terminated by adding boiling water and heating for 10 minutes. Dead enzyme protein was used as a control in all reactions to obtain enzyme reaction products.

[0058] 2. HPLC detection

[0059] The enzyme reaction product was identified by HPLC, and the HPLC detection conditions were as follows: Agela Technologies chromatographic column (Venusil XBPCI8(A), 5 μL, 250mm×4.6mm); flow rate was 1mL / min; injection volume was 5 μL; mobile phase contained 1.5% (v / v) triethylamine acetic acid solut...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention provides a gene CsRHMb capable of efficiently catalyzing the biosynthesis of UDP-rhamnose as well as encoded protein and an engineered strain of the gene CsRHMb. The gene is obtained by separating and cloning fresh tea leaves, the encoded protein is in a reaction system taking UDP-glucose as the substrate and in which NADPH exists, and the protein or the recombinant protein expressed by the engineered strain can efficiently convert UDP-glucose to generate UDP-rhamnose. The invention provides the efficient and safe biosynthesis technology of UDP-rhamnose, the optimal reaction condition of enzyme is optimized through the bioengineering method, and a foundation is provided for realizing the commercial production of UDP-rhamnose.

Description

technical field [0001] The invention relates to the fields of molecular biology and metabolic engineering, in particular to a gene CsRHMb that efficiently catalyzes the biosynthesis of UDP-rhamnose, its encoded protein and its application. Background technique [0002] As a non-alcoholic beverage, tea is widely accepted and favored by people all over the world. In tea tree, polyphenolic compounds mainly exist in fresh tea leaves in the form of glycosylation. Flavonoid glycosides are the main secondary metabolites in tea trees, and polyphenolic compounds are closely related to tea quality and human health care. Among them, flavonols and their derivative products such as flavonol glycosides are important components that determine the quality and quality of tea. UDP-rhamnose, as an important compound of rhamnose activating molecules, not only participates in the plant cell wall synthesis pathway, but also acts as a precursor for the synthesis of flavonol glycosides. For exam...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N15/61C12N9/90C12P19/24C12P19/02
Inventor 高丽萍代新龙黄克依赵贵福赵月石渝凤黎明夏涛
Owner ANHUI AGRICULTURAL UNIVERSITY
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com