High Efficiency Long Insert Mate Pair Library Preparation for NGS Platforms

Next generation DNA sequencing (NGS) instruments produce gigabases per run, but the short read lengths and small size of sequenced fragments result in gaps, misassembled contigs, collapsed repeats and missing sequences, leaving these regions to be finished manually, if at all.  A technology that provides long range sequence linkage from short reads is needed for accurate, economical de novo assembly of genomes. We developed a >90% efficient mate pair library construction technology that incorporates Chimera Codes™ to distinguish true mate pairs from false junctions. NxMate™ NGS libraries were constructed using a reference E. coli strain, Thermus aquaticus and two repeat rich mouse BACs. Without mate pair libraries the BAC and genome assemblies contained numerous unordered contigs. The addition of >90% efficient NxMate data allowed accurate de novo assembly and closing of the BACs and genomes. For comparison, several Illumina Nextera mate pair libraries were constructed and yielded <5% mate pair data. Ion Torrent “mate pair reads without paired end sequencing” permits economical sequence assembly of BAC clones and small genomes, while NxMate libraries for the Illumina platform allows for long range paired end sequencing and assembly of larger genomes.