The protein tag is a polypeptide or protein expressed as a fusion with the target protein using in vitro DNA recombinant technology, which facilitates the expression, detection, tracer and purification of the target protein. Protein tags are usually divided into three categories: genetic tags, in vivo tags and in vitro tags. The most familiar one is genetic tags, such as c-MYC, FLAG, etc. The target genes are cloned into vectors with tags for detection by antibody or fluorescence. At the same time, tags also facilitate protein purification.
Here are some common tags for you.
Flag tag, a peptide recognized by an antibody (DYKDDDDK). The Kozak sequence constructed in the vector makes the fusion protein with FLAG more efficient in the eukaryotic expression system.
l As a fusion expression tag, FLAG usually does not interact with the target protein and affect the function and properties of the target protein.
l The target protein fused with FLAG can be directly performed affinity chromatography through FLAG. This chromatography is non-denaturing purification, which can purify the active fusion protein with high purification efficiency.
l FLAG, a protein recognized by anti-FLAG antibodies. So it is convenient to detect and identify FLAG fusion proteins by Western Blot, ELISA and other methods.
His6-tag, 6 histidines bound by a nickel or cobalt chelate (HHHHHH), which can be inserted at the C-terminus or N-terminus of the target protein. They have two uses as tags, one is to form an epitope to facilitate purification and detection, the other is to form a unique structural feature (binding ligand) to facilitate purification. The side chain of histidine residues has a strong attraction to solid nickel and can be used for immobilized metal chelation chromatography (IMAC) to separate and purify recombinant proteins.
l The molecular weight is small, only ~0.84KD, which generally does not affect the function of the target protein.
l His tag fusion protein is also used for protein-protein and protein-DNA interaction research.
l His tag has relatively low immunogenicity, and the purified protein can be directly injected into animals to prepare antibodies for immunization.
l It is applied to a variety of expression systems with mild purification conditions. And it is also used to construct dual affinity tags with other affinity tags.
GST (Glutathione-S-transferase) tag, a protein that binds to immobilized glutathione, and its natural size is 26KD. There are two reasons for application in prokaryotic expression:
1. It is a highly soluble protein and is used to increase the solubility of foreign proteins.
2. It plays a role in increasing the amount of expression because it is expressed in large quantities in E. coli.
The GST fusion expression system is widely used in the expression of various fusion proteins and can be expressed in host cells such as E. coli and yeast.
C-Myc tag, a small tag containing eleven amino acids (Glu-Gln-Lys-Leu-Ile-Ser-GluGlu-Asp-Leu). These eleven amino acids are expressed as epitopes in different protein frames, but they can still recognize the corresponding antibody.
C-Myc tag has been successfully applied in Western-blot hybridization technology, immunoprecipitation and flow cytometry to detect the expression of recombinant proteins in target cells.
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