Starting material lanes 1–4, yeast lysate, untreated with RNase I; lanes 5–6, immunopurified ribosomes; lanes 7–10, total ribosomes. Numbers to the left of each figure indicate marker sizes in nt. A. RNA before removal of ribosomal RNA. Samples in lanes 2 and 5 are degraded and should be re-prepared; if the problem persists, it is necessary to make another preparation of lysate and ribosomes. Samples in lanes 5–10 have fragmented ribosomal RNA due to incubation of the lysate from which they were prepared with RNase I; this is expected. B. RNA after removal of ribosomal RNA. Samples in lanes 5 and 9 have incomplete removal of ribosomal RNA; these RNAs should be subjected to another round of ribosomal RNA removal. C. Finished libraries after PCR and cleanup. Sample in lane 5 is a library with a broad size range. This library should be prepared again if sequencing results indicate poor quality.