Figure 6.

 Knock-in of EPHB4 mutation in HEK293T cells results in increased mTORC1 activity. (A) Schematic drawing of targeted genome editing at EPHB4 locus using CRISPR/Cas9. (B) Sanger sequence traces of genomic DNA after the knock-in. (C) RT-PCR analysis suggested a small insertion in the knock-in cells compare with the wild-type cells. (D) The four-amino-acid insertion induced by CRISPR/Cas9 confirmed by standard Sanger sequencing of RT-PCR product. (E) Western blot of the wild-type HEK293T cells and EPHB4 mutant cells by gene editing. Cells containing the EPHB4 splice-altering mutation displayed higher p-p70S6K levels than wild-type cells.