Methodology
Plate-based transfection and culturing technique for genetic manipulation of Plasmodium falciparum
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* Corresponding author: Joseph L DeRisi joe@derisilab.ucsf.edu
1 Department of Biochemistry and Biophysics, University of California San Francisco, San Francisco, California, USA
2 Codexis, Inc., 200 Penobscot Drive, Redwood City, California 94063, USA
3 Howard Hughes Medical Institute, University of California San Francisco, San Francisco, California, USA
Malaria Journal 2012, 11:22 doi:10.1186/1475-2875-11-22
Published: 18 January 2012Abstract
Genetic manipulation of malaria parasites remains an inefficient, time-consuming and resource-intensive process. Presented here is a set of methods for 96-well plate-based transfection and culture that improve the efficiency of genetic manipulation of Plasmodium falciparum. Compared to standard protocols plate-based transfection requires 20-fold less DNA, transient transfection efficiency achieved is approximately seven-fold higher, whilst stable transfection success rate is above 90%. Furthermore the utility of this set of protocols to generate a knockout of the PfRH3 pseudogene, screened by whole-cell PCR, is demonstrated. The methods and tools presented here will facilitate genome-scale genetic manipulation of P. falciparum.