Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 102, Issue 45, Pages 16391-16396Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0504679102
Keywords
malaria; mutagenesis; transposon
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Funding
- NIAID NIH HHS [R01AI33656, R01 AI033656, R01 AI048561, R01 AI033656-13, R01AI48561] Funding Source: Medline
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Functional analysis of the Plasmodium falciparum genome is restricted because of the limited ability to genetically manipulate this important human pathogen. We have developed an efficient transposon-mediated insertional mutagenesis method much needed for high-throughput functional genomics of malaria parasites. A drug-selectable marker, human dihydrofolate reductase, added to the lepidopteran transposon piggyBac, transformed parasites by integration into the A falciparum genome in the presence of a transposase-expressing helper plasmid. Multiple integrations occurred at the expected TTAA target sites throughout the genome of the parasite. We were able to transform A faliparum with this piggyBac element at high frequencies, in the range of 10(-3), and obtain stable clones of insertional mutants in a few weeks instead of 6-12 months. Our results show that the piggyBac transposition system can be used as an efficient, random integration tool needed for large-scale, whole-genome mutagenesis of malaria parasites. The availability of such an adaptable genetic tool opens the way for much needed forward genetic approaches to study this lethal human parasite.
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