The work carried out and described in the previous pages was within the prokaryotic bacterial system. Applying CRISPR to the eukaryotic mammalian human system is a different and more complex feat for two primary, related reasons—human cells are protected by two phospholipid bilayers at the plasma membrane and nuclear membrane levels.
Work by Dr. Zhang
Creative work carried out by Dr. Feng Zhang successfully solved the problem in 2013.10 The scientists selected a cell line called 293T cells derived from human embryonic kidney cells. A very complex design involving multiple CRISPR molecules, Cas9 from S. pyogenes together with necessary enzymes, promoters, and nuclear localization signals (NLS), the design successfully expressed CRISPR apparatus in the human cell nucleus. This accomplishment is of huge significance because it opened doors for CRISPR as a gene-editing tool to modify genes of interest in human cells. More details regarding CRISPR-edited pig heart transplantation to the first human will be described in the next section.
10. Ran, F. A., Hsu, P. D., Wright, J., Agarwala, V., Scott, D. A., & Zhang, F. (2013). Genome engineering using the CRISPR-Cas9 system. Nature protocols, 8(11), 2281–2308. https://doi.org/10.1038/nprot.2013.143
