KLF4 K225/229R mutant has a novel NheI restriction site. e) pCS2-KLF4-K225/229R construct diagram. f) NheI / EcoRI Restriction digest of both wild-type and successfully mutated plasmid DNA. g) α-Flag Western blot showing expression of mutant construct in 293T cells. h) Sequencing result of the mutation, mutated residues are highlighted in red.