Tuesday, May 10, 2011

Ribo-Zero rRNA removal from FFPE RNA

One of the key benefits of the Ribo-Zero™ family of rRNA removal products is their ability to remove rRNA from partially degraded total RNA samples. We recently presented data on the performance of Ribo-Zero Kits with formalin-fixed, paraffin-embedded (FFPE) samples at the AACR 2011 Annual Meeting.

RNA from FFPE breast tumor tissue was used as starting material to prepare a mRNA-Seq library. The RNA sample was treated with the Ribo-Zero rRNA Removal Kit (Human/Mouse/Rat) prior to library preparation using the ScriptSeq™ mRNA-Seq Library Preparation Kit (Illumina®-Compatible). Single-lane, 54-nt unidirectional sequencing reads were obtained using an Illumina® GAII sequencer, and sequence alignment was performed by Beijing Genomics Institute (BGI) with both the well-annotated human genome (ENSEMBL56) and in-house built junction sequences using BWA software allowing for four mismatches. The results were compared to those obtained from a library prepared without Ribo-Zero treatment.

(click to enlarge figure)

The Ribo-Zero-treated sample contained 76% of all 49,222 possible expressed genes with 0.24% of the reads being rRNA, compared to 48.8% rRNA reads for the untreated samples. As seen in the figure, Ribo-Zero treatment significantly increases mapped reads for exon, intergenic, and intron transcripts.

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