Figure 2From: Development of the piggyBac transposable system for Plasmodium berghei and its application for random mutagenesis in malaria parasitesLocation of PiggyBac inserts into the genome of P. berghei. A. PiggyBac insertions as shown by FIGE analysis of separated chromosomes hybridized with the pbdhfr/ts probe. This probe recognizes the inserts and the endogenous dhfr/ts gene on chromosome 7 (arrow). Left panel: Inserts in two parent parasite populations (1, 2) after transient transfection of the helper (h) and donor (d) plasmid (h/d ratio: P1 = 1:1; P2 = 1:2; c = control parasites transfected with only donor plasmid). Right panel: insertions in 15 (a to o) parasite subpopulations of P1. B. PiggyBac insertions as shown by FIGE analysis of separated chromosomes hybridized with the pbdhfr/ts probe, which recognizes the endogenous dhfr/ts gene (chromosome 7) and the transposase construct pL1307 in chromosome 5 (arrows). Left panel: Insertions in the three parent parasite populations after transient transfection of the donor (d) plasmid into parasites of mutant TPSama1 that contains transposase integrated into chromosome 5 (amount of d: P3 = 15 μg; P4 = 10 μg; P5 = 5 μg). Right panel: insertions in 10 parasite subpopulations of P5 (a to l). C. Upper panel: WebLogo representation of the sequence of 127 piggyBac insertion sites, showing the TTAA insertion site and 20 bp up- and downstream of the piggyBac 5'ITR and 3'ITR, respectively. Lower panel: WebLogo representation of 20 bp up- and down-stream sequence of 254 randomly chosen TTAA sites in the P. berghei genome. D. Left panel: Chromosomal distribution of the 254 randomly chosen TTAA sites and 124 piggyBac TTAA insertion sites. Right panel: location of the piggyBac inserts (black bars) and 254 random TTAA sites (white bars) in CDS (+introns), within 1 kb 5' or 1 kb 3' to the CDS (designated as 5' UTR and 3 'UTR) or in the intergenic regions (> 1 kb from CDS).Back to article page