Construction and application of genome rearrangement technology of a sugar alcohol-producing yeast strain

A technology of sugar alcohol and recombinant strains, applied in the direction of microorganism-based methods, fungi, and treatment of microorganisms with electricity/wave energy, to achieve low cost, high safety factor, and simple operation

Active Publication Date: 2019-04-30
TIANJIN INST OF IND BIOTECH CHINESE ACADEMY OF SCI
View PDF2 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the high-yield, high-tolerance and other excellent characteristics of industrial strains are jointly determined by a large number of genes, and most of the problems faced by the improvement of industrial microbial strains

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
  • Construction and application of genome rearrangement technology of a sugar alcohol-producing yeast strain
  • Construction and application of genome rearrangement technology of a sugar alcohol-producing yeast strain
  • Construction and application of genome rearrangement technology of a sugar alcohol-producing yeast strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0016] Example 1: Construction and optimization of high-efficiency sugar alcohol colorimetric screening method: Prepare 4, 6, 8, 10, 12, 14, 16 (g / L) D-arabitol standard solutions respectively, and add 20 μl of the standard solutions to Deep well plate, then add 500 μl potassium periodate solution. After mixing, it was left at room temperature for 10 minutes, and then 400 μl of 1% L-rhamnose solution was added to neutralize excess periodate ions. After further mixing, 600 μl of Nash reagent was added and reacted at 63° C. for 20 minutes. After cooling, transfer 200 μl of the reaction solution to a shallow well plate, detect its absorbance at 412 nm, and make a standard curve. According to the above method, take 20 μl of the sample to be tested for detection, and the accumulation of sugar alcohol can be judged according to OD412.

[0017] Polysaccharide alcohols are oxidized by periodate under acidic conditions (pH1.0) to generate formaldehyde, and the residual periodate is n...

Embodiment 2

[0021] Example 2 Construction of a yeast protoplast fusion screening method based on dual fluorescent dye markers: In yeast protoplast fusion, when two yeast strains containing complementary genetic markers are fused, their fusions can grow on selective media. However, many industrial yeast strains, due to the lack of selectable genetic markers, cannot achieve complementary selection of genetic markers [12]. In order to solve this problem, fluorescence activated cell sorting (fluorescence activated cell sorting, FACS) as an effective method is applied to fusion cell screening. In this method, parental strains are labeled with different fluorescent dyes, and fusions containing overlapping fluorescent stains are screened by flow cytometry. This technique is currently used in some applications, but has not yet formed a systematic approach [13-15].

[0022]In the present invention, first prepare 1.0ml of different parental yeast protoplasts, stain with 0.25μM NuclearGreen and 0.1...

Embodiment 3

[0024] Example 3 Application of a new genome rearrangement technology in the improvement of the sugar alcohol-producing strain Pichia anomaly: In order to further effectively improve the yeast sugar alcohol production performance, the strains (U-7, 9 and A-1, 4) is the starting bacterium. Using the new genome rearrangement technology constructed, different high-yield sugar alcohol strains are combined and rearranged to screen strains with excellent performance. The specific process is as follows image 3 . First, the strains were activated and cultured in YPD liquid medium for 12 hours to the logarithmic growth phase, and then the cell wall was removed. After treatment with 2.5% helicase solution at 30°C for 3 hours, the strains were observed by microscope inspection and plate coating. At this time, they had better Protoplast formation rate and regeneration rate. Next, the prepared protoplast cells were labeled with different fluorescent dyes (Nuclear Green and Nuclear Red), ...

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

A novel sugar alcohol-producing yeast strain genome rearrangement technology mainly includes high throughput screening of a sugar alcohol-producing yeast strain on the basis of a color development method and fusion cell high-efficient flow sorting on the basis of bi-fluorescent staining. By means of the technology, high-effective genome rearrangement and screening of alcohol-producing yeast are achieved. The technology is simple, low-cost, high-efficient and practical. Through the technology, high-efficient and quick improvement on natural pichia pastoris being abnormal in sugar alcohol producing can be successfully achieved, thereby increasing the yield of sugar alcohol by more than 34%.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a genome rearrangement technology of a sugar-alcohol-producing yeast strain and the application of the technology in genome transformation of sugar-alcohol-producing Pichia anomala. Background technique [0002] Sugar alcohols refer to the polyols formed after the aldehyde and ketone carbonyl groups of sugars are reduced to hydroxyl groups, with the general formula H(CHOH) n+1 H, which loses its reductibility due to hydrogenation, has a hydroxyl group attached to each carbon atom. They have many unique properties, such as low metabolic energy in the body, can prevent dental caries, and have low insulin metabolic response. Therefore, they are an important class of functional food raw materials. In addition, sugar alcohols also have more applications in the fields of medicine, chemical industry and animal nutrition. In the field of medicine, they are important int...

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
Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/01C12N13/00C12N15/04C12N1/16C12N1/19C12R1/84
Inventor 王钦宏张国强贺鹏
Owner TIANJIN INST OF IND BIOTECH CHINESE ACADEMY OF SCI
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap