Fig. 1. Antisense RNA specifically inhibits ROP2 expression. Analysis of the steady-state levels of protein of ROP2AS clones was done by immunoblot with equal numbers of parasites using a monoclonal antibody T34A7 (Sadak et al., 1988). (A) Because the antibody also recognizes ROP3 and ROP4, each clone conveniently serves as its own internal control for loading and specificity of targeting by antisense RNA. (A) The autoradiogram was over-exposed to demonstrate the faint ROP2 band in the ROP2AS-7 lane. Densitometric scanning of a less intensely exposed autoradiogram demonstrated that ROP2 expression was lowered by 87-92% for ROP2AS-1, ROP2AS-7, ROP2AS-10 and ROP2AS-20, whereas ROP2 expression in the ROP2AS-8 clone was normal, probably reflecting recovery (see Fig. 6). The limited variation among the four repressed clones may be partly fortuitous, but in addition probably reflects the fact that the majority of clones (>80%) were not recovered at all, thereby skewing the results in favor of parasites expressing basal levels of ROP2. (B) ROP2 and NTPase levels are unaffected by expression of antisense HXGPRT construct and irrelevant vector control pminCAT.

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