Uncovering the Complexity of Transcriptomes with RNA-Seq

Figure 2

Library preparation and clonal amplification. Schematic representation of a workflow for library preparation in RNA-Seq experiments on the SOLiD platform. In the figure is depicted a total RNA sample after depletion of rRNA, containing both polyA and non-polyA mRNA, tRNAs, miRNAs and small noncoding RNAs. Ribo-depleted total RNA is fragmented (1), then ligated to specific adaptor sequences (2) and retro-transcribed (3). The resulting cDNA is size selected by gel electrophoresis (4), and cDNAs are PCR amplified (5). Then size distribution is evaluated (6). Emulsion PCR, with one cDNA fragment per bead, is used for the clonal amplification of cDNA libraries (7). Purified and enriched beads are finally deposited onto glass slides (8), ready to be sequenced by ligation.