POLE-LACZ-P286R sequencing

Direct Sequencing of DNA POLE P286R Polymerization Reaction Products to generate the substrate for the direct sequencing of POLE synthesis products, 3 approximately equally spaced DNA primers were hybridized to single strand M13mp2 DNA. Reactions were initiated by the addition of POLE P286R. The double stranded product was then formed into next generation sequencing libraries using a ThruPLEX-FD kit (Rubicon Genomics). Libraries were individually barcoded and pooled. Pooled libraries were sequenced on a single MiSeq lane (Illumina) using a 2X150 paired-end protocol.