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Characterizing cell heterogeneity using PCR fingerprinting of surface multigene families in protozoan parasites

Abstract:

Parasites counteract the action of the immune system and other environmental pressures by modulating and changing the composition of their cell surfaces. Surface multigene protein families are defined not only by highly variable regions in length and/or sequence exposed to the outer space but also by conserved sequences codifying for the signal peptide, hydrophobic C-terminal regions necessary for GPI modifications, as well as conserved UTR regions for mRNA regulation. The method here presented exploits these conserved signatures for characterizing variations in the mRNA expression of clonal cell populations of protozoan parasites using a combination of nested PCR amplification and capillary electrophoresis. With this workflow, in silico gels from isolated cell clones can be generated, thus providing an excellent tool for analyzing cellular heterogeneity in protozoan parasites.