shRNA directed against firefly luciferase and interruption of shRNA by an intron. (A) Schematic representation of the formation of the stem-loop structure through the self-splicing of a group I intron inserted into the loop site. (B) Expression vectors were constructed for shRNAs targeting the "155-173" or "851-875" region of firefly luciferase. The shRNA cassettes were placed under control of the CMV promoter of the pRNA-CMV3.1-Neo vector. The loop portion of the shRNA was from the loop region of hsa-mir-371 with a slight modification. The loop sequences are underlined. The inserted site of the group I intron is illustrated, and the modified nucleotides are shown in red. (C) Schematic representation of the interactions involved in self-splicing including the pairing between the intron and the 5' and 3' exon sequences. These pairings, namely the P1 helix (5'exon-intron pairing) and P10 helix (intron-3'exon pairing), are shown in bold, and the modified nucleotides are shown in red.