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Amplicon Sequencing

Amplicon sequencing has emerged as a reliable and cost-effective ultra-deep targeted, next-generation sequencing (NGS) method to discover and profile known, rare, and novel variants. This highly adaptable approach is utilized in various fields ranging from agriculture and metagenomics to oncology and inherited disease profiling, becoming a standard method of interrogating several targets in multiple samples in a single run. Amplicon sequencing provides an accurate yet economical alternative to whole genome or whole exome sequencing, allowing researchers to focus on genes of interest without the burden of high cost and cumbersome data analysis.

Swift Biosciences’ advanced technology offers an array of pre-designed and customizable Accel-Amplicon™ Panels optimized for multiple sequencing platforms to enable a fast, comprehensive approach to targeted sequencing. Leveraging a combination of unique design and chemistry, the Accel-Amplicon Panels produce overlapping (tiled) amplicons in a single-tube using a rapid 2-hour workflow. This unique molecular biology approach provides powerful solutions for detecting genetic variations and screening clinically-relevant germline and low frequency somatic mutations in putative disease genes using low input DNA from challenging samples types, such as FFPE and cfDNA. A fast and easy single-tube workflow produces the best-in-class performance for on-target percentage and coverage uniformity, enabling low frequency variant discovery and confirmation.

SWIFT PRODUCT LINES COMPATIBLE WITH AMPLICON SEQUENCING:

Accel-Amplicon™ NGS Panels

The Swift advantage:
  • Informative — Interrogate thousands of germline and somatic variants down to 1% in clinically-relevant genes.
  • Streamlined — Easy, single-tube workflow to generate sequence-ready samples in under 2 hours.
  • Complete — Enables manual or automated workflow from content definition through data analysis.
  • Reliable — Uniform coverage and on-target performance from the most challenging and degraded samples, including FFPE and cfDNA.
  • Adaptable — Compatible with Illumina and Ion Torrent sequencing platforms.
  • Flexible — Enables either pre-designed or custom sequencing panels.
Cancer Gene Panels
Rare Disease and Prenatal/NIPT Research Panels
  • Accel-Amplicon CFTR Panel — Screen disease-relevant mutations and variants in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene
Sample-Tracking and Quality Control
  • Accel-Amplicon Sample_ID Panel — Use high minor allele frequency SNPs to identify and track unique molecular signatures by gender and origin, and confirm tumor-normal pairs
Custom NGS Panels
  • Customizable Content — Collaborative design process; large panels 15-1,500 amplicons; pooled and tested using our unique validation process
High Coverage Uniformity Across Sample Types

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The Accel-Amplicon 56G Oncology Panel was used to prepare libraries from 10 ng of input DNA from a variety of sample types. The coverage uniformity, as the percentage of the bases covered at least 20%, 30%, 40%, or 50% of the average depth, was determined across four sample types. The percentage of reads on target was > 95% for all sample types.
Reproducible Variant Calling from Q-Seq HDx™ Quantitative Standards

AP_Amp Seq_Table_Reproduce Variant

The Accel-Amplicon 56G Oncology Panel consistently detected validated variants at the expected frequency in replicates from 10 ng of the Horizon Diagnostics Quantitative Multiplex DNA Reference Standards HD701. The variants were called by LoFreq 2.1.1 (Genome Institute of Singapore) and GATK HaplotypeCaller (Broad Institute). When examining uncommon variants between the 10 replicates, the majority of background variants were present at less than 0.6%. No sporadic variants greater than 0.6% were detected.
Robust Yields and Performance from Clinical Oncology Research Samples

Endometrial study by John Martignetti and Peter Dottino, Icahn School of Medicine at Mount Sinai, for analysis of somatic mutations in 56 oncology-related genes from over 200 samples.

AP_Amp Seq_Table_Edometrial Study

The Accel-Amplicon 56G Oncology Panel was used to assess the germline variant profile of 39 normal samples and 162 tumor samples of varying quality. Following qPCR input quantification using the ALU115 assay, 10 ng of input DNA was used per sample and libraries were sequenced using MiSeq® v2 chemistry. Tumor samples were sequenced to a target depth of 5000X for somatic variant detection while normal samples were sequenced to a target depth of 500X for establishment of a background variant profile for subtraction during variant calling. Bases on target is the percentage of aligned bases on target and coverage uniformity is the percentage of amplicons covered greater than 20% of the mean.