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Gene expression profiling of Leishmania (Leishmania) donovani: overcoming technical variation and exploiting biological variation

Decuypere, S.
Vanaerschot, M.
Rijal, Suman
Yardley, V.
Maes, L.
de Doncker, S.
Dujardin, Jean-Claude
Published in Parasitology. 2008, vol. 135, no. 2, p. 183-94
Abstract Gene expression profiling is increasingly used in the field of infectious diseases for characterization of host, pathogen and the nature of their interaction. The purpose of this study was to develop a robust, standardized method for comparative expression profiling and molecular characterization of Leishmania donovani clinical isolates. The limitations and possibilities associated with expression profiling in intracellular amastigotes and promastigotes were assessed through a series of comparative experiments in which technical and biological parameters were scrutinized. On a technical level, our results show that it is essential to use parasite harvesting procedures that involve minimal disturbance of the parasite's environment in order to 'freeze' gene expression levels instantly; this is particularly a delicate task for intracellular amastigotes and for specific 'sensory' genes. On the biological level, we demonstrate that gene expression levels fluctuate during in vitro development of both intracellular amastigotes and promastigotes. We chose to use expression-curves rather than single, specific, time-point measurements to capture this biological variation. Intracellular amastigote protocols need further refinement, but we describe a first generation tool for high-throughput comparative molecular characterization of patients' isolates, based on the changing expression profiles of promastigotes during in vitro differentiation.
Keywords AnimalsDNA Primers/chemistryGene Expression Profiling/methodsGene Expression Regulation, DevelopmentalGenes, Protozoan/physiologyHumansLeishmania donovani/genetics/growth & development/physiologyLeishmaniasis, Visceral/parasitologyLife Cycle Stages/physiologyPolymerase Chain Reaction/methodsTime Factors
PMID: 17931458
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Research group Groupe Jean-Michel Gaspoz (23)
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DECUYPERE, S. et al. Gene expression profiling of Leishmania (Leishmania) donovani: overcoming technical variation and exploiting biological variation. In: Parasitology, 2008, vol. 135, n° 2, p. 183-94. doi: 10.1017/S0031182007003782 https://archive-ouverte.unige.ch/unige:1881

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Deposited on : 2009-06-04

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