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Targeted Sequencing of Cell-Free DNA with Ion AmpliSeq Technology

By 10.15.2014

Ion World Tour Manchester Presentation: “Targeted sequencing of cell-free DNA using the Ion AmpliSeq Cancer Hotspot and Custom Gene Panels” by Dr. David Guttery, University of Leicester, Leicester UK

The concept of interrogating cell-free DNA from circulating plasma samples has gained popularity,ultrapure-DNA-red.jpg and has been called a ‘liquid biopsy’.  At Ion World Manchester, Dr. David Guttery and colleagues have been researching the use of the Ion PGM™ system with the Ion AmpliSeq™ Cancer Hotspot Panel V2 to detect low levels of circulating DNA.

The amounts vary over time, as free DNA is cleared in the bloodstream relatively quickly. He noted that their yield is typically on the order of 1 to 10 ng per mL, and that DNA extraction would need to take place within two hours of sample collection or stored at -80°C to avoid degradation of the sample.

He presented data that titrated known input amounts into a high amount of background normal control DNA, and detected a minor allele frequency down to 5%, at a 98% sensitivity across 300 previously studied samples.

Also he mentioned they designed a custom Ion AmpliSeq™ panel specifically to interrogate Copy Number Variation for tumor samples, and had data that showed they could accurately call copy number variants down to an input amount of 1ng target DNA per mL equivalent normal background. Dr. Guttery noted that the online Ion AmpliSeq™ Designer was “a really neat tool” in order to specifically design for CNV analysis.

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