Equinox Library Amplification Kits deliver excellent fidelity, uniform sequence coverage, and high library complexity to specifically address the stringent demands of applications such as rare variant detection, circulating cell-free DNA (cfDNA) analysis, single-cell analysis, and hybridization capture. Kits contain a uniquely engineered, ultra-high fidelity DNA polymerase in an optimized hot start PCR mix formulated for high efficiency, low-bias NGS library amplification.
Key Features and Benefits
Ultra-high fidelity amplification that reduces misincorporation events by up to 40%
Reduced overall error rates and facilitation of robust error correction enable rare mutation detection
Effective hot start formulation inhibits both polymerase and exonuclease activities,
important for low-input applications and automated workflows
Efficient library amplification from a wide range of inputs (0.1 pg to 500 ng) and GC content (15% to 85%)
Highly uniform sequence coverage optimizes sequencing economy
Compatibility with paragmagnetic beads ensures robust performance in hybrid capture workflows.
Low-frequency variant detection NGS assays, including those utilizing challenging samples such as FFPE and cfDNA
Hybridization capture workflows
Whole genome sequencing
- Amplicon sequencing
- ChIP-Seq, ATAC-Seq, and associated epigenetic applications
- Illumina and non-Illumina sample preparation workflows
Key Performance Data
Ultra-high fidelity system enables high sensitivity applications
The Equinox Amplification Master Mix contains a proprietary proofreading polymerase optimized for ultra-high fidelity amplification, delivering a 40% reduction in overall polymerase error rate in comparison to KAPA HiFi HotStart ReadyMix. This enables sensitive variant detection by helping to minimize overall error rates and reducing false variant calls. The significant reduction in C>T substitutions is particularly important as this mutation is associated with the spontaneous deamination of methylated cytosine to uracil, and is also one of the most common mutation types in cancers.
Low-bias amplification delivers uniform UMI family coverage
Unique Molecular Indices (UMIs; also known as molecular barcodes) are added to sequencing libraries prior to PCR amplification to enable accurate bioinformatic identification of PCR duplicates and improve variant calling in low-input applications. However, biased amplification, where a small number of molecules are preferentially amplified at the expense of others, results in uneven representation across UMI families and can generate a large number of singleton UMI bins, which are not compatible with error correction. A significant amount of additional sequencing may be required to attain requisite coverage of low abundance sequences. Equinox Library Amplification Kits enable uniform amplification across UMI families, supporting coverage for >75% of all read families (and >90% of read families with GC content from 25 – 75%) within 3X of the mean family depth.
Effective hot start formulation safeguards automated performance
The Equinox Amplification Master Mix is formulated with a highly effective hot start mechanism, which strongly inhibits both the 5’→ 3’ polymerase and 3’ → 5’ exonuclease activities of the enzyme. Such inhibition mitigates primer and low input sample degradation, as well as artifacts resulting from nonspecific amplification, improving sensitivity in low-input applications and better facilitating automated library construction.
High-efficiency amplification limits bias and artifacts
The Equinox Library Amplification Kit delivers high efficiency amplification, making it possible to limit amplification bias and artifacts by reducing the number of cycles needed to generate the desired yield for downstream steps. Amplification efficiency remains robust when targeting yields greater than 1000 ng, minimizing bias for applications demanding high yields, such as hybridization capture workflows.
Uniform amplification improves sequencing economy
Equinox Library Amplification Kits introduce minimal amplification bias, even in the presence of paramagnetic beads. This simplifies the workflow, as well as ensures compatibility with hybridization capture workflows. Additionally, Equinox delivers even coverage uniformity across complex genomes which can reduce the amount of sequencing needed to achieve desired coverage depths.