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Pull-down assay experiments using Tagged protein purification kits
Introduction of Pull-down assay experiments using Tagged protein purification kits
Proteins function by interacting with various other proteins. To reveal the function of a protein, it is important to identifiy other proteins which interact to the protein.
Protocol
Experiment conditions
Exampe 1: Pull-down assay of an endogenous protein
p62/SQSTM1 directly interacts with LC3, which is localized on autophagosome membrane, and is degradated by autophagic-lysosome pathway. Interactions of p62/SQSTM1 with LC3 was confirmed by pull-down assay using the cell lysate of the HA-LC3 transfectant.
HA tagged Protein PURIFICATION KIT (MBL Code No. 3320)
Example 2: Pull-down assay of forcibly expressed proteins
Beclin 1, which is essential for early autophagosome formation, is reported to make complex with various proteins such as UBRAG and Vps34 and regulate the processes of autophagosome/lysosome fusion (Autophagosome maturation) and endocytosis. Using HA, c-Myc, and His-tagged protein purification kits, pull-down assay of Beclin1 and UVRAG is performed
*Samples: HA tagged Beclin1/Myc-His tagged UVRAG transfectant.
HA tagged Protein PURIFICATION KIT (MBL Code No. 3320)
c-Myc tagged Protein MILD PURIFICATION KIT Ver.2 (MBL Code No. 3305)
His tagged Protein PURIFICATION KIT (MBL Code No. 3310)
Tag-protein Purification Kit & Gel