Antisense sRNAs are detected for the silenced EhROM1 gene in G3:ROM(KD) parasites. Top, strand-specific oligonucleotide probes to the EhROM1 gene (arrows in black are probes with Northern blots pictured; arrows in gray are the probes without Northern blots pictured but mentioned under “Results” as data not shown). Four sense probes for detecting antisense sRNAs are shown as RS-86 (ROM1 sense probe starting at position −86), RS1 (ROM1 sense probe starting at position 1), RS168 (ROM1 sense probe starting at position 168), and RS471 (ROM1 sense probe starting at position 471). The trigger region used for psAP-2 cloning is shown below (between positions −86 to 447). Bottom, Northern blot analysis detects antisense sRNAs for EhROM1 in G3:ROM(KD) parasites. A Northern blot (loaded with 50 μg of sRNA-enriched sample from E. histolytica trophozoites of strain HM-1:IMSS, Rahman, G3, and G3:ROM(KD)) was probed with oligonucleotide probe RS-86 to detect antisense sRNAs; the blot was stripped and probed with ap-a probe S1. Methylene blue staining is shown as a loading control.