Epitope Tagging
1.
Epitope
tagging is used for localization and purification of protein.
2.
Using standard
engineering techniques, a short peptide tag can be added to protein interest.
3.
It is
used to study protein-protein or DNA-protein interaction.
4.
If a tag
itself is an antigen determinant or epitope it can be targeted by an appropriate
commercially available antibody.
5.
The antibody,
suitably labelled can be used to determine the location of protein in cell to
purify it by immunoprecipitation of affinity chromatography.
6.
In an immunoprecipitation
antibody directed against the epitope tag re added to a solution as antibody
protein complex.
7.
Richard
young and his co-workers
originally identified 10 polypeptide that are authentic polymerase 2 sub at
least tightly bond contained.
8.
The method
they used is called epitope tagging, in which they attach a small foreign
epitope to one of yeast poly II subunit (RPBS)
9.
Then they
introduce this gene into yeast cell, lacking a functional RPB3 gene.
labelled the protein with either S35 or P32 and use as Ab
directed against the foreign epitope to precipitate whole enzyme.
10.
After immunoprecipitation,
they separated the labelled polypeptide of precipitated protein by SDS page and
detected them by autoradiography.
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