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Pegfp c3 mst2

Manufactured by Addgene

The PEGFP C3-Mst2 is a plasmid vector that contains the coding sequence for the Mst2 (Mammalian Sterile 20-like 2) protein fused to the green fluorescent protein (GFP) sequence. This vector allows for the expression and visualization of the Mst2 protein in transfected cells.

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2 protocols using pegfp c3 mst2

1

Overexpression and RNAi of YAP/TEAD Pathway

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For overexpression experiments, YAP–HaloTag CRISPR knock-in U-2 OS cells were transfected with pEGFP-C3-Lats1 (Addgene plasmid # 19053) or pEGFP C3-Mst2 (Addgene plasmid # 19056), both gifts from Marius Sudol, pRK5-Myc-Fus-R495X,68 (link) gift from Jiou Wang, or EGFP-TAZ (made in the Cai lab) using Lipofectamine 3000 Transfection Reagent (cat. no. L3000015), for 16 h. For RNAi experiments, a mixture of Lats1 siRNA (Thermo Fisher, Silencer Select s17393) and Lats2 siRNA (Thermo Fisher, Silencer Select s25503) was used at a final concentration of 10 nM, or the scrambled negative control siRNA was used at a final concentration of 20 nM (Thermo Fisher, AM4611), and were transfected into cells using the Lipofectamine RNAiMAX transfection reagent (Thermo Fisher, cat. no. 13778075) for 48 h. For siTEAD1 experiments, the TEAD1 siRNA (Thermo Fisher, Silencer Select s13962) or scrambled negative control siRNA was used at a final concentration of 10 nM (Thermo Fisher, AM4611), and were transfected into cells using the Lipofectamine RNAiMAX transfection reagent (Thermo Fisher, cat. no. 13778075) for 48 h, after which cells were replated for live cell imaging and RT-qPCR.
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2

Fascin and MST2 Modulation in Cellular Processes

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Fascin targeting sequence GAAGAAGCAGATCTGGA was inserted to lenti-CRISPR V2 vector according to previous report [22 (link)]. Wild-type fascin expression plasmid was generated by inserting full-length fascin cDNA into pLncx2 retrovirus vector at BglII and EcoRI sites. Fascin-S39E mutant was prepared using Quick Change Mutagenesis kit. Plpcx-flag-MST2 was generated by inserting Flag-MST2 cDNA into plpcx vector at XhoI and EcoRI sites. Full length and truncated fragments (1-313AA and 314-491AA) of MST2 were inserted to PET28a vector at BamHI and XhoI sites.
Plasmids of pEGFP-C3-MST2 (Plasmid #19056), pLenti-EF-FH-TAZ-ires-blast (Plasmid #52083) and pLenti-EF-FH-TAZS89A-ires-blast (Plasmid #52084) were obtained from Addgene.
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