Detecting CRISPR/Cas gene doping

CRISPR/Cas changes DNA and is considered "gene doping" and is banned by the World Anti-Doping Agency, an independent international organization. A sufficient method to detect CRISPR/Cas gene editing needs to be developed, however. So, Mario Thevis and colleagues wanted to see whether they could identify the protein most likely to be used in this type of doping, Cas9 from the bacteria Streptococcus pyogenes (SpCas9), in human plasma samples and in mouse models.

The team spiked the SpCas9 protein into human plasma, then isolated and cut it to analyse by mass spectrometry . The researchers found that they could identify unique components of the SpCas9 protein from the complex plasma matrix. In another experiment, inactivated SpCas9, which can regulate gene expression without altering DNA, was spiked into human plasma samples. With a slight modification, the method allowed the team to purify and detect the inactive form. Finally, the team injected mice with SpCas9 and showed that their concentrations peaked in circulating blood after 2 hours and could be detected up to 8 hours after administration into muscle tissue. The researchers say that although much work still needs to be done, this is an initial step toward a test to pinpoint athletes trying to gain an unfair advantage.