Chinese hamster ovary (CHO) cells are the workhorse mammalian cell line for producing complex biopharmaceuticals. The plasticity of CHO cell lines is advantageous to the production of recombinant proteins such as monoclonal antibodies (mAbs). However, this plasticity causes cell line instability due to gene or even chromosome rearrangements. Genome instability can result in transgene copy loss and decreased protein titer in long- term culture. Cell line instability is a key challenge for the field to overcome for a more rapid and low-cost complex biopharmaceutical production. DNA double-stranded breaks (DSBs) can cause DNA mutations that reduce therapeutic protein production, threaten genome integrity, and can result in cell death. In CHO cell lines, some DNA repair genes were mutated from the conserved region of other mammalians including Chinese hamsters. The mutated genes can be restored by a site- specific modification to study its effect on cell line stability. Mutated genes with single nucleotide mutations were chosen to be the target genes for prime editing modification. I successfully accomplished gene disruption and knockout using CRISPR Cas9 or Cas12a plasmids. I established a stable gene modification workflow in the Sandoval lab. The gene knockout clones can be used as a control group to compare with the gene-restored group. I demonstrated the first-time prime editing method in CHO cells, and this expanded the CHO genome engineering toolbox. I developed a five-step method to increase the prime editing efficiency from 0% to 13.9% using PE2 and combined pegRNA-knot-PE3-D2 plasmid. I developed a robust protein purification and glycosylation analysis method to compare glycan profiles for dynamic pH and limited feeding bioreactor conditions for CHO- produced VRC01 protein. The limited feeding conditions had a more desired glycan profile. Future directions for studying the effect of restored DNA repair genes in CHO cell lines were proposed. Critical quality attributes (CQA) in long-term shake flask culture can be measured to determine the gene-restored cell line stability. The more stable gene- restored cell lines can be studied in bioreactors. Restoring multiple DNA repair genes could further increase cell line stability.