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. 2007 May 30;2(5):e472. doi: 10.1371/journal.pone.0000472

Figure 6. Small RNA guides induce Rnt1p cleavage in vivo.

Figure 6

(A) Mapping the RNA-directed cleavage of U2 from living yeast cells. Yeast cells were electroporated with 2 nmols of EU2dT or mutant RNA guides and total RNA was extracted between 10 minutes and 2 hours after electroporation. The primer PE-U2-Br was annealed with the RNA samples and extended on U2 snRNA. The corresponding DNA sequence is shown on the left. (B) Relative levels of cleaved U2 snRNA after electroporation were established using RNA extracted in A and loaded on 1.2% agarose-formaldehyde gel, transferred to a nylon membrane and hybridized with probes specific for nucleotides 7 to 29 into mature U2 or against the 25S rRNA to serve as internal control and visualized by Phosphor Imager. The intensity of each band was quantified using Image Quant 5.0. The levels of U2 snRNA were normalized against the 25S rRNA and the electroporation efficiency (53±8%), and were plotted as a function of time after electroporation with EU2dT.