Gene Knockout

(1) Short fragment removal: Guide RNAs target introns at both sides of exon 2. The number of bases in exon 2 is not a multiple of 3, which can cause a frame-shift mutation.

(2) Frame-shift mutation: Guide RNAs target the exon. The base number of the deleted exon is not a multiple of 3, which can cause a frame-shift mutation.

(3) Large fragment removal: Complete removal of the coding sequence to achieve the gene knockout.

Gene Point Mutation

The gRNAs, Cas9 and the Donor vector are co-transferred into the target cells by nucleofection. After the gRNA/Cas9 complex causes a DNA double-strand break at the targeting site, the Donor fragment carrying the designed point mutation is used as a template for homologous recombination repair (HDR), thus being able to introduce the designed point mutation into the targeting site of the genome.

Gene fragment Knockin

(1) Knockin of a fusion protein fragment (such as an EGFP tag) at a specific locus.

(2) Knockout of a specific gene plus knockin of an exogenous gene.

Gene Overexpression

The vector expressing the target gene will be transferred into the selected cell lines by either lentivirus transduction or nucleofection. After drug screening, the stable cell lines will be selected.

Gene Knockdown

The knockdown vector will be transferred into the selected cell lines by either lentivirus transduction or nucleofection. After drug screening, the stable cell lines will be selected.

Work Flow

Project Report and Final Product

• Positive clone screening report
• Cell cryopreservation tube

Types of Gene Editing Cell Lines (successfully generated)