The Small RNA-Seq Library Prep Kit provides a protocol for generating small RNA libraries for Illumina sequencing directly from total RNA or enriched small RNA.
Multiplexing of up to 96 samples is possible with complimentary i7 indexes provided in the kit. This allows you to pool more samples per sequencing lane and perform cost-efficient experiments on the platforms of different scale – from bench top to high throughput instruments.
Gel-free User-Friendly and Fast Protocol
Lexogen’s Small RNA-Seq kit offers a time saving protocol that can be completed within 5 hours, and requires just about 1 hour of your hands-on time. The final library does not need to go through gel purification. Quick and convenient magnetic bead-based purification can optionally be performed to remove adapters dimers for some demanding RNA inputs.
The protocol exhibits exceptional reproducibility and correlation across samples with the different concentrations (Figure 1).
Figure 1. a) Dilution series of purified plasma RNA in Small RNA-Seq shows high inter-replicate correlation Reads per Million (RPM) across concentrations tested (60 pg and 600 pg). 1 b) Dilution series of purified plasma RNA in Small RNA-Seq shows high correlation RPM across concentrations tested (60 pg and 600 pg). As an example, only data for 60 pg and 600 pg are shown.
The protocol allows for detection of a higher number of microRNAs than other workflows. The difference is especially considerable for lower RNA inputs (Figure 2).
Figure 2. Total number of miRNAs detected across 4 different protocols. Dilution series (6 pg, 60 pg, and 600 pg) of purified plasma RNA used for library prep from different vendors were performed. The obtained libraries were sequenced at equal molarity at ~ 1.5 – 2M Total Raw Reads per sample. Lexogen’s Small RNA-Seq Kit showed much higher numbers of detected miRNAs at ≥ 5 Raw Reads across all concentrations tested.
Optimized for RNA from Challenging Sources
The high sensitivity of the protocol makes it very well suited for challenging, low content RNA sources, such as liquid biopsies (plasma, serum, and urine), including exosomes. The protocol has also been tested on RNA from cells and tissues. The kit can be used for inputs from 100 ng – 1,000 ng of cellular total RNA or 50 pg – 1,000 ng enriched small RNA including plasma, serum, and urine.
Lexogen’s SPLIT RNA Extraction Kit offers the opportunity to extract NGS-grade quality total RNA or small and large RNA fractions from the same sample. SPLIT is highly suitable for purification of total RNA and enrichment for small RNA for use with the Small RNA-Seq Library Prep Kit. Thus, the combination of these two protocols offers you the complete solution for small RNA sample preparation for sequencing on any of the Illumina platform.