Characterizing PARP7 Interactions with NF-κB

Cos-1 cells were transfected with 0.5 µg of p50 cFlag pcDNA3 or RelA cFlag pcDNA3 alone or along with either 1 µg of pEGFP-PARP7 or 0.8 µg of pEGFP-PARP7H532A using Lipofectamine™ 2000 (Invitrogen). After 24 h, cell lysis and co-immunoprecipitation was carried out as previously described [22 (link)]. The antibodies used for detection with Western blotting were anti-poly/mono-ADP-ribose (Cell Signaling Technology; E6F6A), anti-GFP (rabbit) (Abcam, Cambridge, United Kingdom; ab290), anti-GFP (mouse) (Clontech Laboratories, Mountain View, CA, USA; JL-8), anti-FLAG (rabbit) (Sigma Aldrich; F7425), and anti-FLAG (mouse) (Sigma-Aldrich; M2).

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Rasmussen M., Alvik K., Kannen V., Olafsen N.E., Erlingsson L.A., Grimaldi G., Takaoka A., Grant D.M, & Matthews J. (2023). Loss of PARP7 Increases Type I Interferon Signaling in EO771 Breast Cancer Cells and Prevents Mammary Tumor Growth by Increasing Antitumor Immunity. Cancers, 15(14), 3689.

Publication 2023

Lipofectamine™ 2000 anti-poly/mono-ADP-ribose anti-GFP (rabbit) anti-GFP (mouse) anti-FLAG (rabbit) anti-FLAG (mouse)

Antibodies Cell Co immunoprecipitation Cos 1 cells Lipofectamine 2000 Mouse Poly adp ribose Rabbit Rela

Corresponding Organization : University of Toronto

Other organizations : University of Bradford, Hokkaido University

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Variable analysis

independent variables

0.5 µg of p50 cFlag pcDNA3
0.5 µg of RelA cFlag pcDNA3
1 µg of pEGFP-PARP7
0.8 µg of pEGFP-PARP7H532A

dependent variables

Protein interactions detected through co-immunoprecipitation and Western blotting

control variables

Cos-1 cells used as the cell line
Lipofectamine 2000 used for transfection
Cell lysis and co-immunoprecipitation carried out as previously described in [22]

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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