What is the purification method of the protein purification system

What is the purification method of the protein purification system? It is necessary to know the coding DNA sequence of the purified protein, to see which cells or tissues are overexpressed in the target gene, and to design gene primers to amplify the arf of the target DNA fragment. This is the so-called acquisition of target gene fragments.

Construction of expression vector: In the prokaryotic or eukaryotic expression vector with the characteristics of the obtained gene expression itself, the main problem of this step is to construct the plasmid and the gene of interest, and the expression system. The prokaryotic expression time is short, the cost is low, and a large amount of expression is a priority; the gene is not expressed in E. coli, and the problem occurs in codon optimization. In consideration of better activity and purification of the protein, the researchers chose to express in Pichi yeast. Successful expression of codon optimization is important.

What is the purification method of the protein purification system:

1. Precipitation.

2. Electrophoresis: A charged protein is higher or lower than its isoelectric point and can be moved to the negative electrode or the positive electrode of the electric field in the electric field. Support film electrophoresis, electrophoresis, etc.

3. Dialysis: A method that uses two dialysis bags to separate large molecules from proteins and small molecular organic compounds.

4. Chromatography: Ion exchange chromatography uses the free properties of proteins. Under a specific pH, the charges and properties of proteins are different, and they can be separated by ion exchange chromatography. In anion exchange chromatography, proteins with low negative power are eluted first. Molecular sieves, also known as gel filtration. Small proteins enter the pores and stay in them for a long time. Large proteins cannot enter the pores and flow out directly.

5. What is the purification method of the protein purification system? Ultracentrifugation: Protein purification can be used to determine the molecular weight and can be used as the protein. The formation of proteins with different densities is separated.