USE OF ZINC FINGER NUCLEASE TECHNOLOGY TO IMPROVE GENETIC MANIPULATION OF THE CAUSATIVE AGENTS OF CHAGAS’ DISEASE AND AFRICAN SLEEPING SICKNESS
Genetic manipulation of the parasites Trypanosoma brucei and Trypanosoma cruzi depends on integration via homologous recombination. Zinc finger nucleases (ZFN) are artificial DNA-binding proteins that can be engineered to recognize and cleave specific sequences. It has recently been reported that the creation of double-strand breaks in the genome of T. brucei improves transformation efficiencies. As proof of concept we will test the ability of ZFN targeted to a synthetic sequence to improve stable transformation of both T. cruzi and T. brucei. We will also examine the efficiency with which ZFN targeted against T. cruzi genes can produce null mutants and test how readily the phenotype can be rescued by integrating plasmid libraries into the synthetic site recognized by the first ZFN. A long-term goal is that complementation with plasmid libraries can be performed routinely, allowing genes causing specific phenotypes to be identified in both parasites.
Prof. Isabel RODITI
Prof. Santuza TEIXEIRA