A Method for Rapid Genetic Integration into Plasmodium falciparum Utilizing Mycobacteriophage Bxb1 Integrase

Genetic manipulation of the human malaria parasite Plasmodium falciparum has presented substantial challenges for research efforts aimed at better understanding the complex biology of this highly virulent organism. The development of methods to perform gene disruption, allelic replacement or transgene expression has provided important insights into the function of parasite genes. However, genomic integration studies have been hindered by low transfection and recombination efficiencies, and are complicated by the propensity of this parasite to maintain episomal replicating plasmids. We have developed a fast and efficient site-specific system of integrative recombination into the P. falciparum genome, which is catalyzed by the mycobacteriophage Bxb1 serine integrase. This system has the advantage of providing greater genetic and phenotypic homogeneity within transgenic lines as compared to earlier methods based on episomal replication of plasmids. Herein, we present this methodology.