Step 2b: CRISPR/Cas9-mediated gene knock-in
This step is similar to step 2a. The differences are as follows:
1. A donor plasmid must be constructed in order to insert a cassette, such as EGFP, mCherry or another Tag sequence, into the genome by homologous recombination.
2. Southern blot and DNA sequencing will be used to verify the cassette insertion.
3. It takes one more month to make a knock-in cell line than to make a regular gene knockout line.