EQUIPLEX™ NORMALIZATION KITS
Balanced by design
Cas9-mediated normalization for consistent, scalable NGS workflows
Product Overview
Library quantification and normalization remain a persistent bottleneck for NGS labs, introducing variability at a critical point in the workflow. This step is prone to error: double dipping, skipped wells, and inaccurate serial dilutions of high-concentration libraries often lead to uneven representation or “jackpotting” in pooled samples. The result is inefficient sequencing at best, or rework and resequencing at worst, which disrupts processes, increases costs and time to result, and adds operational complexity.
EquiPlex™ is designed to address these challenges directly, leveraging Cas9 in a fundamentally different way than its genome editing roots — not to cut, but as a programmable, stoichiometric binding tool that enables more consistent, reliable normalization to support smooth, scalable sequencing workflows.
Applications
- →RNA sequencing, including 3' sequencing
- →Methylation sequencing
- →DNA sequencing, including ChIP-seq and metagenomics
- →Targeted sequencing (post-capture)
- →Single cell sequencing (non-tagmentation workflows)
- →Amplicon sequencing
Designed for amplified libraries built with TruSeq®-style adapters, including libraries moving directly to sequencing (post-hybrid capture).
Key Features and Benefits
Cas9-mediated normalization
EquiPlex repurposes Cas9 as a stoichiometric binding tool, tagging a fixed number of library molecules for bead-based capture. Both kit configurations deliver consistent normalization in under one hour without degrading or single-stranding libraries. The workflow is fully automatable, with no off-deck steps required.
Figure 1. Rapid, automatable normalization of NGS libraries. Both configurations — the Singleplex Kit, which normalizes libraries individually, and the Multiplex Kit, which normalizes libraries in pools — tag a fixed number of molecules with a Cas9-mediated binding reagent and use bead-based capture to select for bound molecules. The entire process takes less than one hour manually and is fully automatable, with no manual interventions or off-deck steps required.
A better way to normalize
Compared to quantify-and-dilute approaches, EquiPlex is the only normalization solution that combines fast turnaround, full automation, and low readcount variability without manual calculations, specialized equipment, or off-deck handling.
| Criteria | EquiPlex Normalization | Quantify-and-Dilute Methods | |||
|---|---|---|---|---|---|
| qPCR | MiSeq®/iSeq® | Fluorimetric¹ | Electrophoretic² | ||
| Fast turnaround time | +++ | – | – | ++ | – |
| Automatability, including no off-deck handling | +++ | – | – | ++ | – |
| Low readcount variability | +++ | ++ | +++ | + | + |
| No manual calculations, dilutions, or variable volume pipetting | +++ | – | – | – | – |
| No additional or specialized equipment | +++ | – | – | – | – |
¹ Examples: Qubit®, Picogreen® ² Examples: TapeStation®, Fragment Analyzer®
More uniform pools for more efficient sequencing
Uniform library representation means more confident multiplexing, less top-off sequencing, and more output from every flow cell. Across a 500-fold input range into library prep, EquiPlex delivers consistently low read count variability without affecting insert size, GC bias, or data quality, and entirely eliminates jackpotting to ensure every sample receives its fair share of reads.
Figure 2. Eliminate jackpotting and maintain library quality across a 500-fold input range. (A) 96 DNA libraries generated from NA12878 (1–500 ng; 8 replicates per input mass) were normalized in 96-plex using EquiPlex or manual methods. EquiPlex achieves a CV of 10%, while the manual control shows a CV of 34%, including two dropouts and an incidence of jackpotting. (B) GC coverage profiles for 500 ng input libraries normalized by either method are nearly identical, confirming no impact of normalization method on data quality.
Normalization without data compromise
EquiPlex fits seamlessly into RNA-seq workflows, delivering uniform read distributions across a 400-fold input range (2.5–1000 ng into mRNA library prep) without compromising data integrity. Gene expression profiles show excellent concordance between EquiPlex and manual normalization, with no measurable effect on percent alignment, gene body coverage, or unique genes detected.
Figure 3. Uniform read distribution and preserved gene expression across broad input ranges. (A) mRNA libraries spanning a 40-fold concentration range (5–200 nM) were normalized in a single 96-plex batch using the EquiPlex Normalization Kit (Multiplex); normalized read contributions were ~1% per library across all input concentrations. (B) Correlation analysis of gene expression data from 96-plex pools shows excellent concordance between EquiPlex and manual normalization across a 400-fold RNA input range (all R values >0.99).
Built for flexibility
The EquiPlex Normalization Kit (Multiplex) supports pool sizes from 8 to 96 libraries per batch, with reagents included for up to 96 total libraries per kit. Different pool sizes can be combined within a single run, giving you the flexibility to match any experimental design — with no requirement for spatial adjacency or multiples of eight.
Technical Specifications
| Input | 5–200 nM in 10 µL (TruSeq®-style adapters) |
| Kit formats | Singleplex and Multiplex |
| Batch size | 8 to 96 libraries per run |
| Total workflow time | Under one hour (Bind → Bead Capture → Wash → Elute → Collect) |
| Storage | Part 1 at −20°C ± 5°C; Part 2 at 2–8°C |
| Shelf life | ≥6 months (24 rxn) or ≥12 months (>24 rxn) |
| Read count variability | CV of ~10% vs. 34% for manual quantify-and-dilute (96-plex, DNA) |
Ordering Information
| Product | 24 RXN | 96 RXN |
|---|---|---|
EquiPlex Normalization Kit (Singleplex) Incl. reagents for library binding, bead capture, bead washing, and elution | 7BK0016-024 | 7BK0016-096 |
EquiPlex Normalization Kit (Multiplex) Incl. reagents for library binding, bead capture, bead washing, and elution | 7BK0017-024 | 7BK0017-096 |
For larger pack sizes, contact us. For Research Use Only. Not for use in diagnostic procedures.