Fig. 1. Overview of the IMPRESS technology.

During combined digestion with four MSREs, unmethylated recognition sites are digested, whereas methylated CpG sites, blocking the MSREs, remain intact. Subsequently, methylated CpG sites are captured by hybridization of the smMIP binding arms. Both the selected CpG sites and reference sites are targeted by specifically designed smMIPs, referred to as CpG smMIPs and reference smMIPs, respectively. The insert gap is extended and ligated to form a circular fragment. All linear fragments are degraded by an exonuclease treatment and the circular fragments are amplified by PCR with universal primers binding the common smMIP backbone. Eventually, PCR products are pooled, purified and sequenced simultaneously. Created with BioRender.com.