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. 2007 Feb 14;35(5):1612–1623. doi: 10.1093/nar/gkm031

Figure 7.

Figure 7.

Inverse RT-PCR analysis for detection of B.c.I4 lariat and other circular molecules. A; schematic drawing illustrating the inverse PCR strategy and the location of primers. B; in vivo analysis; gel electrophoresis of nested RT-PCR products obtained from total RNA isolated from B. cereus ATCC 10987 cultures. C; in vitro analysis; gel electrophoresis of RT-PCR products obtained from in vitro self-splicing of B.c.I4 wild-type (WT). Sequencing chromatograms of selected inverse RT-PCR products are shown to the right. The black, grey and white arrows indicate full-length circles, molecules linked within the extra 56-nt segment, and lariats branched in the bulged A:899, respectively. In B and C, lane M shows the size marker, pBR322 DNA digested with MspI (New England Biolabs); in B, lane NC shows a negative control done without reverse-transcriptase. Primers I4A_right/I4A_right_nested and I4B_left were used in B; I4A_right and I4B_left_lariat were used in C (Table 1). Samples were separated on a 3.5% NuSieve GTG agarose gel (Cambrex).