siRNA Wee1

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	siRNA concentration 40 nM Incubate the FuGENE 6 Transfection Reagent/medium mixture for 5 minutes at room temperature  Incubate transfection mixture for 15 min at room temperature and add to cells and shake for 10-30 seconds.  Incubate cells for 96h.

Protocol tips
siRNA concentration 40 nM Incubate the FuGENE 6 Transfection Reagent/medium mixture for 5 minutes at room temperature  Incubate transfection mixture for 15 min at room temperature and add to cells and shake for 10-30 seconds.  Incubate cells for 96h.

Publication protocol

MDA-MB-231 and SKOV3 cells were grown on glass coverslips in 6-well plates. If indicated, cells were treated with MK-1775 (500 nM) in the absence or presence of SU-9516 (1 µM). At 14 d after treatment, cells were fixed in 4% (wt/vol) paraformaldehyde in PBS. Subsequently, cells were stained with rat anti-CD44 (BioLegend 103002) combined with Alexa488-conjugated goat anti-rat secondary antibody and counterstained with DAPI. Images were obtained using a Leica DM6000B microscope equipped with a 10× objective (Leica HC PL FLUOTAR 10×/0.30) and LAS-AF software (Leica). In total 100–500 cells were scored per condition. Alternatively, cells were grown on glass coverslips and transfected with 40 nM Wee1 siRNA (#1 WEE1HSS111337 and #2 WEE1HSS111338; Invitrogen) or “medium GC duplex” control siRNA (Life Technologies 12935–300). At 96 h after siRNA transfection, cells were fixed and stained as described above.

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Manufacturer protocol

Download the product protocol from Thermo Fisher Scientific for siRNA Wee1 below.

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