An improved approach to mate-paired library preparation for Illumina sequencing

High quality data from mate-pair libraries provides long range sequence linkage across the genome, which is crucial for de novo assembly and structural-variant detection. Current commercial methods available for the construction of such libraries have differing limitations and are often linked to a single sequencing platform in a kit format, which may not be cost effective. We present an alternative mate-paired protocol, demonstrated using Illumina sequencing platforms, combining the specificity of hybridisation and ligation, to circularise fragments with high yield. An adapter sequence is incorporated between the junction site of the mate pairs, the length of which is evenly controlled by nick translation. We present a comparison of results from 3 Kb E. coli and Plasmodium falciparum 3D7 mate-pairs made with our protocol, alongside commercial mate-pair methods. Furthermore, we present the results of a set of 3 and 6 Kb mate-pair libraries from seven different mouse strains made with our mate-pair protocol to demonstrate its reliability and robustness.