Tagmentation method for preparing DNA for NGS sequencing from as little as 20pg input DNA amount, as developed by the Systems Biology Laboratory UK.

New sequencing technologies can address diverse biomedical questions but are limited by a minimum required DNA input of typically 1 microgram. We describe how sequencing libraries can be reproducibly created from 20pg input DNA using a modified transpososome-mediated fragmentation technique. Resulting libraries incorporate in-line barcoding which facilitates sample multiplexes that can be sequenced using Illumina platforms with the manufacturer's sequencing primer. We demonstrate this technique by providing deep coverage sequence of the E. coli K-12 genome that shows equivalent target coverage to a 1mg input library prepared using standard Illumina methods. Reducing template quantity does, however, increase the proportion of duplicate reads and enriches coverage in low GC regions. This finding was confirmed with low coverage re-sequencing of mouse from 20pg gDNA input, equivalent to ~7x haploid genomes, where ~0.4x coverage of mapped fragments were recovered. Application of this new method now allows genomic studies from low mass samples and routine preparation of sequencing libraries from enrichment procedures.