Sanger Sequencing

The gold standard method to identify induced mutations at the target locus entails amplifying the target region by PCR, cloning the amplicon into a vector, followed by Sanger sequencing of the amplicon directly. Each vector should carry only one gene copy, which generates a clean trace in the chromatogram after sequencing. This approach reveals both the frequency and type of mutation at the target locus.  It is commonly used to perform screening on clonal cell lines to confirm and validate the desired mutation.​