Gene deletion from Plasmodium falciparum using FLP and Cre recombinases: implications for applied site-specific recombination.
The ability to manipulate the genome and induce site-specific recombination using either Flippase (FLP) or Cre recombinase has been useful in many systems including Plasmodium berghei for specific deletion events or to obtain conditional gene expression. To test whether these recombinases are active in Plasmodium falciparum we constructed gene knockouts that contain sequences recognised as templates for site-specific recombination. We tested the ability of FLP and Cre recombinases, expressed conditionally in P. falciparum, to mediate deletion of the human dihydrofolate reductase (hdhfr) drug resistance gene. We show that Cre recombinase is capable of efficient removal of hdhfr by site-specific recombination. In contrast, FLP recombinase is very inefficient, even at the optimum temperature of 30°C for this enzyme. These results demonstrate that Cre recombinase can be utilised in P. falciparum for deletion of specific sequences such as drug resistance genes. This can be exploited for recycling of drug resistance cassettes and for the design of specific recombination events in P. falciparum.
O'Neill
Matthew T
MT
The Walter and Eliza Hall Institute of Medical Research, 3052 Melbourne, Australia.
Phuong
Thuan
T
Healer
Julie
J
Richard
Dave
D
Cowman
Alan F
AF
D004254:DNA Nucleotidyltransferases / Q000378:metabolism*; D004351:Drug Resistance; D017353:Gene Deletion; D055786:Gene Knockout Techniques / Q000379:methods*; D005827:Genetics, Microbial / Q000379:methods*; D006801:Humans; D019426:Integrases / Q000378:metabolism*; D010453:Peptide Synthases / Q000235:genetics; D010963:Plasmodium falciparum / Q000235:genetics*; D011995:Recombination, Genetic
C045073:Cre recombinase; D004254:DNA Nucleotidyltransferases; C045119:FLP recombinase; D019426:Integrases; D010453:Peptide Synthases; C020041:dihydrofolate synthetase