Figure 1.
Microarray gene expression data from schizont and ring /uninucleate trophozoite stage parasites from a primary P. yoelii infection.
Yir gene expression signals (log2) in schizont (blue) and ring /uninucleate trophozoite (red) stage parasites for each gene are paired and shown as bars, for the complete set of yir genes analyzed in this experiment. Positive gene expression values, above the 90th percentile cut-off in the starter population, are shown. Data points for all yir (226) genes displaying positive gene expression values, in the samples analysed, are included in the graph (see also Supplementary Table S1 for gene identity (ID)). Cut-off levels for genes transcribed at high (yellow), medium (green) and low (purple) levels are indicated by shading. Expression of several non-yir genes (Pcfam homologues (a), multiple banded antigen (b), ribosomal subunit protein L23 (c), β-tubulin (d)) is shown for comparison.
Figure 2.
Group assignment and chromosomal location of yir genes transcribed in blood stage infection of P. yoelii.
The proportion of transcribed yir genes (a) and transcribed subtelomeric yir genes (b) in each group is compared to the proportion expected from their representation in the P. yoelii genome. Inset: Proportion of yir genes of each group in the genome (adapted from Fonager et al, 2007).
Figure 3.
RT-PCR on single micromanipulated schizonts and uninucleate trophozoites.
a) Location of the conserved yir primer pairs in a genomic setting. f1 (forward primer 1)/r1 (reverse primer 1) and f2/r2 pairs are shown with black arrows, while the yir gene structure is shown in blue, sizes are indicated below the gene in bp. b) Single cell RT-PCR on six micromanipulated schizonts (+ lanes 2–12). c) Single cell RT-PCR on seven micromanipulated uninucleate trophozoites (+ lanes 2–14). Expected product sizes were 500–600 bp, controls were reactions from which reverse transcriptase enzyme was omitted (− lanes 3–13 for schizonts and 3–15 for trophozoites). Marker was 100 bp DNA ladder.
Table 1.
Yir transcripts found in individual micromanipulated erythrocytes infected with P. yoelii at the schizont or ring/trophozoite stage of development.
Figure 4.
Microarray analysis showing changes in yir transcription patterns in cloned parasites.
a) Transcription pattern of yir genes in the starter population of P. yoelii ring/uninucleate trophozoite stages (R, in red) and schizont stage (S, in blue) parasites. Each gene is indicated as a bar, expression signal is as defined in figure 1; b) Comparison of the transcription profile of 578 yir genes used as the repertoire of transcribed yir genes derived from infections initiated with a single infected erythrocyte (by limiting dilution). The repertoire of transcribed yir genes in schizont and ring/uninucleate trophozoite stage parasites was determined after ∼10 days of infection. The microarray fingerprints represent the repertoire of genes transcribed from individual mice infected with a single infected erythrocyte obtained from the initial infection (top array and figure 1). [See also Supplementary Table S1, part b for transcribed genes common to all samples].
Figure 5.
Quantitation of selected genes by qRT-PCR in stage-separated parasites.
Comparison of transcription levels of the yir genes PY05826, PY03177, PY04021, PY01966 and PY02298 in stage separated parasites (Rings and uninucleate trophozoites (R) are in red and Schizonts (S) are in blue) isolated from four parasite clones (Clones 1–4), expressed as number of copies transcribed /1×10−9 g β-tubulin.