Eureka AIR delivers breakthrough ideas for toughest innovation challenges, trusted by R&D personnel around the world.

Fusion expression method for green fluorescent protein

A technology of green fluorescent protein and fusion expression, which is applied in the field of fusion expression of green fluorescent protein, can solve the problem of expensive kits, and achieve the effects of avoiding sample loss, good reproducibility and less reagent consumption.

Inactive Publication Date: 2018-06-29
UNIV OF SCI & TECH LIAONING
View PDF6 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, many biochemical reagent manufacturers have special kits for sale, but the price of purchasing kits is relatively expensive

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
  • Fusion expression method for green fluorescent protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] A method for fusion expression of green fluorescent protein, comprising enzyme digestion, connection, transfer and sequencing, specifically comprising the following steps:

[0056] 1. Green fluorescent protein (Green Fluorescent protein, GFP) digestion method

[0057] 1) Digestion of green fluorescent protein with EcoRI enzyme

[0058] Add green fluorescent protein, sterilized water, buffer solution and EcoRI enzyme into a centrifuge tube, heat in a water bath at 35-40°C for 1-1.5 hours, then add solutions PCI, CIA, and sodium acetate in sequence for centrifugal concentration , to remove excess lipids and sugar impurities in the solution, centrifugation conditions: centrifugation temperature 3-5°C, centrifuge speed 10000-14000rpm, centrifugation time 5-10min, to obtain green fluorescent protein DNA with EcoRI enzyme cleavage site , stored in TE solution;

[0059] 2) Digestion of green fluorescent protein with HindIII enzyme

[0060] Heat the green fluorescent protein...

Embodiment 2

[0082] A method for fusion expression of green fluorescent protein, comprising enzyme digestion, connection, transfer and sequencing, specifically comprising the following steps:

[0083] 1. Green fluorescent protein (Green Fluorescent protein, GFP) digestion method

[0084] 1) Digestion of green fluorescent protein with EcoRI enzyme

[0085] Add green fluorescent protein, sterilized water, buffer solution and EcoRI enzyme into a centrifuge tube, heat in a water bath at 35-40°C for 1-1.5 hours, then add solutions PCI, CIA, and sodium acetate in sequence for centrifugal concentration , to remove excess lipids and sugar impurities in the solution, centrifugation conditions: centrifugation temperature 3-5°C, centrifuge speed 10000-14000rpm, centrifugation time 5-10min, to obtain green fluorescent protein DNA with EcoRI enzyme cleavage site , stored in TE solution;

[0086] 2) Digestion of green fluorescent protein with HindIII enzyme

[0087] Heat the green fluorescent protein...

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 relates to a fusion expression method for green fluorescent protein. The method includes enzyme digestion, ligation, transformation and sequencing. In the method, elution is carried outwith PCI, CIA and NaAc, and impurities in the reaction system are removed through gradient centrifugation; then the target DNA fragment and a carrier fragment are respectively subjected to CIAP treatment so that terminals of the target DNA fragment and the carrier fragment turn into cohesive terminals, thus improving ligation successful rate. The method reduces influences on sequencing accuracy due to impurities in the sequencing, and is high in purity of the DNA. The method has simple steps and good repeatability, is low in reagent consumption and reduced in experiment cost, and has equal electrophoresis effect as that in kit operation.

Description

technical field [0001] The invention relates to DNA recombination and operation in genetic engineering, in particular to a fusion expression method of green fluorescent protein. Background technique [0002] Genetic engineering (genetic engineering), also known as gene splicing technology and DNA recombination technology, is based on molecular genetics and modern methods of molecular biology and microbiology. Hybrid DNA molecules are constructed in vitro, and then introduced into living cells to change the original genetic characteristics of organisms, obtain new varieties, and produce new products. Genetic engineering technology provides a powerful means for the study of gene structure and function. [0003] Patent 201710232127 discloses a preparation method of green fluorescent protein-coupled agarose, and obtains gene products by PCR; 201610942279.4 discloses an improved green fluorescent protein expression vector and its construction method. site, PCR and enzyme digest...

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 Applications(China)
IPC IPC(8): C12N15/63C12N15/66C07K14/435
CPCC07K14/43595C12N15/63C12N15/66
Inventor 郝晓亮高云全艳玲王勇费爱萍方志刚姜俊羽马诗文
Owner UNIV OF SCI & TECH LIAONING
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
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