A scalable pipeline for highly effective genetic modification of a malaria parasite

Pfander, C. et al. (2011) A scalable pipeline for highly effective genetic modification of a malaria parasite. Nature Methods, 8(12), pp. 1078-1082. (doi: 10.1038/nmeth.1742) (PMID:22020067) (PMCID:PMC3431185)

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Abstract

In malaria parasites, the systematic experimental validation of drug and vaccine targets by reverse genetics is constrained by the inefficiency of homologous recombination and by the difficulty of manipulating adenine and thymine (A+T)-rich DNA of most Plasmodium species in Escherichia coli. We overcame these roadblocks by creating a high-integrity library of Plasmodium berghei genomic DNA (>77% A+T content) in a bacteriophage N15–based vector that can be modified efficiently using the lambda Red method of recombineering. We built a pipeline for generating P. berghei genetic modification vectors at genome scale in serial liquid cultures on 96-well plates. Vectors have long homology arms, which increase recombination frequency up to tenfold over conventional designs. The feasibility of efficient genetic modification at scale will stimulate collaborative, genome-wide knockout and tagging programs for P. berghei.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Otto, Dr Thomas and Billker, Dr Oliver
Authors: Pfander, C., Anar, B., Schwach, F., Otto, T. D., Brochet, M., Volkmann, K., Quail, M. A., Pain, A., Rosen, B., Skarnes, W., Rayner, J. C., and Billker, O.
College/School:College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
Journal Name:Nature Methods
Publisher:Nature Publishing Group
ISSN:1548-7091
ISSN (Online):1548-7105
Published Online:23 October 2011

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