Ultrasensitive mutation detection in liquid biopsies using SiMSen-Seq

Abstract
Detection of cell-free DNA in liquid biopsies offers great poten- tial for use in non-invasive diagnostics and to monitor treatment. However, the allele frequencies are often low in liquid biopsies and challenging to analyze, since standard next-generation sequencing suffers from insufficient sensitivity and digital PCR is not suitable to analyze multiple allele variants. Hence, we have developed SiMSen- Seq: “Simple, Multiplexed, PCR-based barcoding of DNA for Sensitive mutation detection using Sequencing” that allows allele frequen- cies <0.1% to be detected, using several kilobases of DNA. Several barcoding strategies have been reported, but all require long and complex library preparation protocols. SiMSen-Seq was developed to generate barcoded libraries with minimal DNA input (<5ng), flexible target selection and a very simple (∼3 h) library construc- tion protocol. SiMSen-Seq allows detection of variant alleles with easy customization of library content and a protocol that can be implemented in any molecular biology laboratory. In this presen- tation we will discuss assay development and bioinformatics that are used in SiMSen-Seq. In addition, we show data from several applications, focusing on cancer. http://dx.doi.org/10.1016/j.bdq.2017.02.019