Figure 2.

Different lengths and position of the trigger to the luciferase-coding region results in reporter gene silencing. (A) Schematic of constructs used in this assay. (B) Different lengths of the EHI_197520 trigger were fused upstream of the luciferase-coding region. A 48-bp trigger does not silence luciferase, the 102-bp and 75-bp triggers significantly decreased luciferase expression and the 132-bp trigger resulted in complete silencing. (C) Evaluation of the fusion constructs where a trigger was fused either upstream or downstream of the luciferase-coding region. The first 132 bp of EHI_197520 trigger results in luciferase reporter silencing regardless of whether the trigger is fused up- or downstream of the luciferase gene. The last 132 bp of EHI_197520 trigger does not result in luciferase reporter silencing, regardless of whether the trigger is fused up- or downstream of the luciferase gene. The last 450 bp of EHI_197520 trigger results in luciferase reporter silencing when the trigger is fused upstream of the luciferase gene. All constructs with trigger luciferase are expressed well in the Rahman strain, where no AS sRNAs are present (data not shown).