Click to view: PLOS publication
Adapter dimers form when the 5΄ and 3΄ adapters self-ligate without the target library insert. Consequently, amplification of this smaller adapter dimer side product often out-competes amplification of the tagged library. Our modified adapters block adapter-adapter ligation thus improving the adapter dimer to tagged library ratio. This leads to significantly higher quality sequencing data.
Adapters Reduce Adapter Dimers While
Maintaining Mapped Reads
Human total brain RNA was prepped using either TruSeq® Small RNA Sample Prep Kit or CleanTag™ Ligation
Kit for Small RNA Library Prep.
CleanTag™ Produces Superior Results with Samples Containing Low miRNA Abundance, such as Patient Blood and Plasma Samples
Human plasma samples contain a low abundance of small RNAs and have traditionally been difficult to sequence. In collaboration with Hudson Alpha Institute for Biotechnology, we show that the CleanTag™ Ligation Kit for Small RNA Library Prep efficiently tags small RNA and produces significantly more valuable reads when compared to a previously published protocol optimized for adapter dimer reduction.
Adaptor-Dimer Reduction in microRNA-seq library Sequence Read Disposition. 1 uL RNA isolated from 1 mL
of human plasma (~20ng) was used to generate microRNA-seq libraries using the protocol published by Vigneault, et al. (2012)
and the CleanTag™ Small RNA Protocol. The two protocols were performed separately and combined onto one gel separation step,
and microRNA bands (~140bp) were excised, processed and sequenced with 50bp single end protocol on the Illumina MiSeq Platform (total ~15 Million Reads). CleanTag™ modied adapters were diluted 1:4 and 15 PCR cycles were performed.
CleanTag™ Eliminates Need for Gel Purification
1000 ng Human total brain RNA input. AMPure® XP purified. Analyzed by BioAnalyzer®.
CleanTag™ Renders Small RNA-Seq Fully Automatable
The size of the adapter dimer and tagged library are similar in length, thus requiring a gel size selection step to enrich the libraries for the desired product. Because of this, a fully automated workflow was not possible. Now, with TriLink's CleanTag™ technology, there is no for need gel purification, thus allowing full automation of small RNA-seq for the first time.