PTPRR interactome detection using PUP-IT (pupylation-based interaction tagging) Mass Spectrometry

ABSTRACT: To further stably express PTPRR (WT)-PafA or PTPRR (DA)-PafA in iPUP OVCAR5 cell, we subcloned PTPRR-WT or DA, respectively, into the PafA-IRES-EGFP plasmid. Each plasmid was packed into a lentivirus and then transduced into iPUP OVACR5 cells for 48 h. GFP-positive cells were sorted by flow cytometry. The expression of PTPRR (WT)-PafA and PTPRR (DA)-PafA was confirmed by western blotting analysis. PTPRR (WT)-PafA or PTPRR (DA)-PafA expressed iPUP OVCAR5 cells were grown to a cell density of about 75% on 10 cm dishes. We followed the protocol established previous. To prepare PUP-IT samples for mass spectrometry analysis, including doxycycline induction, biotin labeling, cell lysis, streptavidin magnetic beads pull-down, trypsin digestion, and peptide cleaning.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Ovarian Serous Adenocarcinoma Cell Line, Germinal Epithelium Of Ovary

DISEASE(S): Ovarian Carcinoma

SUBMITTER: Yuetong Wang  

LAB HEAD: Gaofeng Fan

PROVIDER: PXD015352 | Pride | 2019-10-30

REPOSITORIES: Pride