AUTOMATED miRNA LIBRARY PREP WITH UDIs FOR REDUCED BIAS AND SUPERIOR DATA QUALITY
NEXTFLEX® Small RNA-seq v3 Automation Kit with UDIs for Illumina® NovaSeq®, MiSeq®, and HiSeq® 2000/2500 Sequencers
- Greater discovery/detection rate reduces sequencing costs
- Randomized adapter technology reduces ligation bias and increases accuracy
- As little as 200 ng input starting material for complete automation
- Barcoded primer sets allows multiplexing of up to 192 samples
- Unique dual index barcoded primer set mitigates index hopping on the Illumina® NovaSeq®, MiSeq®, and HiSeq® 2000/2500 platforms
- Individually sequence-validated indices ensure confidence in purity
- Automated kit for the PerkinElmer® Sciclone® G3 NGS/NGSx and the Zephyr® G3 NGS workstation saves time and resources
UDI Barcodes 1-96, 96 rxns
UDI Barcodes 97-192, 96 rxns
UDI Barcodes 193-288, 96 rxns
UDI Barcodes 289-384, 96 rxns
Small RNA Library Prep with Randomized Adapters for Reduced Sequencing Bias and Unique Dual Indexes to Mitigate Index Hopping
The NEXTFLEX® small RNA-seq v3 automation kit with UDIs uses the same popular chemistry of the NEXTFLEX small RNA-seq kit v3, which incorporates technologies that reduce small RNA ligation bias, increase ligation efficiency, reduce adapter-dimer formation, and build in a new dimension of data quality, integrity, and accuracy with the integration of Unique Dual Indexes (UDIs) for Illumina® sequencing on the MiSeq®, HiSeq® 2000/2500, and NovaSeq® platforms. The addition of UDI barcodes for small RNA sequencing allows confident multiplexing of up to 192 samples, all while mitigating the risk of index hopping and spread of signal that can occur on a patterned flow cell. The purity of each index sequence is validated by sequencing for high confidence in the resulting sequencing data quality.
The NEXTFLEX® small RNA-seq v3 automation kit with UDIs is compatible with cell-free RNA, such as RNA isolated from plasma. Users who wish to deplete the abundant tRNA fragments and Y RNA fragments found in many types of cell-free RNA should use the NEXTFLEX® tRNA/YRNA blocker (not included in kit). If NEXTFLEX® tRNA/YRNA blockers are of interest, please contact us at [email protected]
Completely Automated Small RNA-seq Library Prep
The NEXTFLEX® small RNA-seq v3 automation kit with UDIs has been designed for automation on the PerkinElmer® Sciclone® G3 NGS/NGSx and Zephyr® G3 NGS workstations. Components of the kit have been optimized and validated on these liquid handling platforms, and the UDI barcoded primers are conveniently arrayed in plated format for easy automation.
Enabling Unprecedented High-Level Multiplexing for Small RNA Sequencing Efficiency
96 barcoded primers (barcodes 1 – 96 or barcodes 97-192) are included in the 96 reaction NEXTFLEX® small RNA-seq v3 automation kits with UDIs. For Illumina® sequencing on the MiSeq®, HiSeq® 2000/2500, and NovaSeq® platforms, the barcoded primers behave as unique dual indexes (UDIs), while when used on the Illumina® MiniSeq®, HiSeq® 3000/4000, HiSeqX®, and NextSeq® platforms, the barcoded primers behave as single index barcodes.
Barcoded primers are plated in column format.
For more information contact [email protected].
More miRNA Discovery by Blocking tRNA/YRNA Fragments
Including the NEXTFLEX® tRNA/YRNA blockers into you small RNA-sequencing workflow can help minimize YRNA and tRNA fragments from making their way into your library, which consequently increases miRNA reads. tRNA and YRNA fragments, though they may have unexplored roles as potential biomarkers, are abundant in sample types like biofluids. For researchers, however, who are primarily interested in other small RNA species like miRNAs, reads lost to these abundant tRNA and YRNA fragments in their sample will inevitably lead to a lower number of reads mapping to their RNA species of interest. Coupling the NEXTFLEX® tRNA/YRNA blockers with the NEXTFLEX® small RNA sequencing chemistry and data security of UDIs allows users to maximize the patented and patent-pending technology for ligase-associated bias reduction, which enables superior miRNA discovery for the same sequencing depth.
- NEXTFLEX® 3’ 4N Adenylated Adapter
- NEXTFLEX® 3’ Ligation Buffer
- NEXTFLEX® 3’ Ligation Enzyme Mix
- NEXTFLEX® Adapter Depletion Solution
- NEXTFLEX® Adapter Inactivation Buffer
- NEXTFLEX® Adapter Inactivation Enzyme
- NEXTFLEX® 5’ 4N Adapter
- NEXTFLEX® 5’ Ligation Buffer
- NEXTFLEX® 5’ Ligation Enzyme Mix
- M-MuLV Reverse Transcriptase
- NEXTFLEX® RT Buffer
- NEXTFLEX® UDI Barcoded Primer 1 – 96 or 97 – 192*
- NEXTFLEX® Small RNA PCR Master Mix
- microRNA Control
- Resuspension Buffer
- Nuclease-free Water
- NEXTFLEX® Adapter Depletion Solution
- NEXTFLEX® Cleanup Beads
* Barcoded primers are plated in column format.
REQUIRED MATERIALS NOT PROVIDED
- Consult with your PerkinElmer® Field Application Scientist or email
SEQUENCES & INDICES
- Sequences of NEXTFLEX Small RNA-Seq UDI Barcode Indexes – Excel / PDF
- Contact [email protected] for instructions for installing NEXTFLEX dual index barcode indices in Illumina® Experiment Manager
- Download the Sage® Pippin Prep® Protocol
- Download the Small RNA Trimming Instructions
- Download the No Size Selection Protocol
ON-DEMAND GENOMEWEB WEBINAR
- Increasing Ligation Efficiency and Discovery of miRNAs for Small RNA NGS Sequencing Library Prep with Plant Samples
- Reduced-Bias Small RNA Library Preparation with Gel-Free or Low-Input Options
- PAG XXIV Poster: Advances in Small RNA Library Preparation Allow Combination of Bias Reduction with Gel-free or Low Input Protocols