CRISPR editing of Topoisomerase II alpha in Trypanosoma brucei

As part of a project seeking modes of action of a novel class of compound that appear to inhibit topoisomerase IIalpha in Trypanosoma brucei we initially selected for mutants resistant to the drug. A number of point mutations were found in the TopoIIa gene and we therefore introduced one of these (F540L) to a cas9 expressing T. brucei line to determine whether editing in this mutation could induce resistance. The edited parasites did acquire resistance to the novel compounds and the work is currently being written for publication.