40 kb Long-Span, Mate-Pair NGS Library Version 2.0

Next generation sequencing (NGS) technologies can rapidly and economically produce a draft genome of an organism de novo. However, the quality of the draft data is seldom more than 80% complete with >10e5 contigs for large genomes, which is insufficient for many applications. Most contigs begin and end with a repeat with existing library construction technologies. Sequence data that is closer to 95% finished with the unambiguous order and placement of genes would have the greatest utility for scientific and commercial research. New molecular tools that bridge the gaps between massively parallel short read sequencing technologies (35-1,500 bases) and the need for large scaffolds (>100,000 bases) to accurately assemble complex repeat rich genomes are needed.  We have successfully developed 40 kb mate-pair NGS libraries by designing and constructing a novel pNGS fosmid system. Our results show that ~70% pNGS fosmid paired-end sequences can be obtained by Illumina sequencing, which is significantly better than existing long-span mate-pair systems. We will introduce version 2.0 of the 40kb long-span, mate-pair NGS fosmid system including an Ion Torrent platform version, and other improvements. We will present data for closing microbial genomes and scaffolding eukaryotic genomes.