Ribosomal RNA Depletion

Enrich whole transcriptome RNA by depleting ribosomal RNA (rRNA)

Our understanding of transcriptome biology is undergoing a revolution. The regulation of expression and the types and functions of RNA are far more complex than was previously thought. Enrichment of whole transcriptome RNA by depleting ribosomal RNA (rRNA) species using our Thermo Scientific RiboMinus technology has the potential to enhance discovery using gene expression microarrays, RNA-Seq, and other methods. This technology can facilitate our understanding of the role of the transcriptome in both normal physiological and pathological processes.

Ribosomal RNA

Ribosomal RNA is a type of non-coding RNA which is the primary component of ribosomes, essential to all cells. Ribosomal RNA is transcribed from ribosomal DNA (rDNA) and then bound to ribosomal proteins to form small and large ribosome subunits. rRNA is the physical and mechanical actor of the ribosome that forces transfer RNA (tRNA) and messenger RNA (mRNA) to process and translate the latter into proteins.[1] Ribosomal RNA is the predominant form of RNA found in most cells; it makes up >80% of cellular RNA despite never being translated into proteins itself.

Ribosomal RNA depletion

Ribosomal depletion is a critical method in transcriptomics that allows for efficient detection of functionally relevant coding as well as non-coding transcripts through removal of highly abundant rRNA species. Use of oligo dT primer to capture the polyadenylated 3′ end of the transcripts and isolate mRNA is routine in many RNA sequencing preparations; however this method lacks the ability to handle degraded samples where most of the RNA is separated from the 3′ tail, or to isolate non-polyadenylated transcripts such as lncRNAs. Ribosomal removal methods address these issues by directly depleting the rRNA while leaving other transcripts intact.

Selecting RNA species of interest

Figure 2. Selecting RNA species of interest. A - Complete total RNA content in eukaryote cell, B - selection of mRNA content in a eukaryote cell by cell oligo dT through Dynabeads mRNA direct Micro kit, C - whole transcriptome content through Ribominus Eukaryote V2.

The process of depleting RNA

In our rRNA removal kits, rRNA is captured by complimentary oligonucleotides that are coupled to paramagnetic beads, after which the bound rRNA is precipitated and removed from the reaction. Using magnetic beads is considered the most efficient way to get high quality RNA.

Good data vs bad data: How rRNA % depletion translates to Next-Generation Sequencing % mapped reads

How removal efficiency of rRNA impacts downstream analysis such as NGS
Figure 3. How removal efficiency of rRNA impacts downstream analysis such as NGS.
Species rRNA Size (kb)
Human 18S  1.9
28S  5
Mouse 18S  1.9
28S  4.7
Drosophila 18S  2
28S  4.1*
Tobacco Leaf 16S  1.5
18S  1.9
23S  2.9
25S  3.7
Yeast 18S  2
26S  3.8
E. coli 16S  1.5
23S  2.9
Xenopus 18S  4
28S  1.8

*Drosophila 28S ribosomal RNA is processed into 2 fragments that migrate in a similar manner to the 18S rRNA.

Ordering information