Plate-based transfection and culturing technique for genetic manipulation of Plasmodium falciparum

• Published in: Malaria journal Vol. 11; no. 1; p. 22
• Main Authors: Caro, Florence, Miller, Mathew G, DeRisi, Joseph L
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 18.01.2012; BioMed Central; BMC
• Subjects: 96-well plate; Automation; Care and treatment; Control; Experiments; Gene expression; Genes; Genetic aspects; Genetic engineering; Genetics, Microbial - methods; Genomes; Genomics; Health aspects; Humans; Malaria; Methodology; Methods; Molecular Biology - methods; Parasites; Physiological aspects; Plasmids; Plasmodium falciparum; Plasmodium falciparum - genetics; Prognosis; Southern blot analysis; Studies; transfection; Transfection - methods
• ISSN: 1475-2875; 1475-2875
• DOI: 10.1186/1475-2875-11-22

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.