RealSeq®-AC is a novel proprietary method for preparing small-RNA sequencing libraries that nearly eliminates incorporation bias in Next-Generation-Sequencing (NSG). Commonly used sequencing library preparations lead to underestimation of the abundance of most miRNAs, some by as much as 10,000-fold.
This under-representation can obscure the presence of some RNAs, including ones that could be useful biomarkers if they were detected accurately. Accurate quantification of microRNAs (miRNA) and other small RNAs is important for understanding their biology and for developing new biomarkers and therapeutic targets.
Most bias stems from sequence-dependent variability in the enzymatic ligation reactions that attach the two adapters to the 3’ and 5’ ends of the miRNAs /small RNAs during preparation of sequencing libraries. By using a novel single adapter and circularization, RealSeq®-AC greatly reduces library preparation bias. Shown below are bias results comparing RealSeq®-AC with a commonly used 2-adapter protocol.
RealSeq®-AC reduces sequencing bias in known problematic miRNAs
RealSeq®-AC reduces incorporation bias compared to other small RNA library preparation kits
RealSeq®- workflow is efficient and gel-free
RealSeq® - detects more miRNAs in total RNA than other miRNA library preparation kits
|Kit I||Kit N||Kit B||Kit T||RealSeq®-AC|
|miRNAs with >5 reads||404||452||412||324||500|
|miRNAs with >10 reads||328||365||352||239||385|
The RealSeq® platform is being expanded to include:
low input samples (single cell level)
targeted RNA- seq
highly fragmented, tumor-derived cell-free DNA (cf-DNA).
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