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An RNA-Seq Strategy to Detect the Complete Coding and Non-Coding Transcriptome Including Full-Length Imprinted Macro ncRNAs

Figure 5

Ribo-depleted RNA-Seq detects macro ncRNAs more efficiently than polyA RNA-Seq.

(A) UCSC genome browser screen shot as in Figure 4A of the Airn macro ncRNA gene. The Cloonan et al. EB polyA RNA-Seq (grey, top), the RiboMinus CCE and Ribo-Zero CCE RNA-Seq data (black, bottom) are shown. Black asterisks mark the signals from the protein-coding mRNA Igf2r and grey asterisks mark the position of a pseudogene expressed from chr.15 [41]. Note that the ncRNA Airn is 118 kb in length (red arrow, extends outside the region shown) and overlaps exons 2 and 1 protein-coding Igf2r gene (black arrow, extends outside the region shown) in antisense orientation. Therefore Airn and Igf2r signals are visible in the CCE data that has no strand-specific information. For Cloonan et al., strand-specific information was available and Igf2r signals are visible on the negative strand (black asterisks, top) whereas only a low amount of signals are visible on the positive strand expressing Airn. (B) As in A showing the functional 83 kb Kcnq1ot1 macro ncRNA (red arrow). (C) As in B showing Mortazavi et al. adult mouse brain polyA RNA-Seq (grey, top), the RiboMinus FH (black, middle) and Ribo-Zero FH RNA-Seq data (black, bottom). (D) As in C showing an annotated RefSeq ncRNA of unknown function. Signals higher than indicated by the scale on the x-axis were cut off. Note that the differences in the read numbers between RiboMinus FH and Ribo-Zero FH reflect the increased number of uniquely aligned tags (see Table S1).

Figure 5