Western Blotting: Protein Analysis

Western blotting was done as previously described (61 (link)) using Hybond PVDF membranes (GE Healthcare) and the following antibodies: GINS3 (Abcam, AB177515), GINS1 (Abcam, AB183524), FLAG (MilliporeSigma, F1804), V5 (Bio-Rad, MCA1360GA), HA (12CA5; Abcam, AB1424), GAPDH (Santa Cruz Biotechnology Inc., sc-25778), and actin (Abcam, AB1801). Bands of interest were quantified by densitometry using ImageJ software (NIH).
For details of immunoprecipitation and immunolabeling procedures, see Supplemental Methods.

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McQuaid M.E., Ahmed K., Tran S., Rousseau J., Shaheen R., Kernohan K.D., Yuki K.E., Grover P., Dreseris E.S., Ahmed S., Dupuis L., Stimec J., Shago M., Al-Hassnan Z.N., Tremblay R., Maass P.G., Wilson M.D., Grunebaum E., Boycott K.M., Boisvert F.M., Maddirevula S., Faqeih E.A., Almanjomi F., Khan Z.U., Alkuraya F.S., Campeau P.M., Kannu P., Campos E.I, & Wurtele H. (2022). Hypomorphic GINS3 variants alter DNA replication and cause Meier-Gorlin syndrome. JCI Insight, 7(10), e155648.

Publication 2022

Hybond PVDF membranes FLAG GAPDH actin

Actin Antibodies Densitometry Gapdh Immunolabeling procedures Immunoprecipitation Membranes Pvdf

Corresponding Organization : Université de Montréal

Other organizations : King Faisal Specialist Hospital & Research Centre, Agricultural Research Institute of Ontario, Newborn Screening Ontario, Université de Sherbrooke, King Fahd Medical City, Alfaisal University

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Variable analysis

independent variables

Antibodies used for Western blotting: GINS3 (Abcam, AB177515), GINS1 (Abcam, AB183524), FLAG (MilliporeSigma, F1804), V5 (Bio-Rad, MCA1360GA), HA (12CA5; Abcam, AB1424), GAPDH (Santa Cruz Biotechnology Inc., sc-25778), and actin (Abcam, AB1801)

dependent variables

Protein expression levels quantified by densitometry using ImageJ software (NIH)

control variables

Western blotting protocol as previously described (61)
Hybond PVDF membranes (GE Healthcare) used for Western blotting

controls

No positive or negative controls were explicitly mentioned in the provided information.

Annotations

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