Library preparation for the major next generation sequencing platforms requires the ligation of specific adaptor oligos to fragments of the DNA to be sequenced. First, DNA is fragmented to the optimal length determined by the downstream platform. Because DNA fragmentation does not result in homogeneous, blunt-ended fragments, end repair is needed to ensure that each molecule is free of overhangs, and contains 5’ phosphate and 3’ hydroxyl groups. Libraries to be used in blunt-ended adaptor ligation, including Ion Torrent™ or SOLiD™ 4 library construction, can be used directly in the ligation step. For Illumina® libraries and some libraries intended for the 454™ platform, incorporation of a non-templated deoxyadenosine 5’-monophosphate (dAMP) onto the 3’ end of blunted DNA fragments, a process known as dA-tailing, is necessary. dA-tails prevent concatamer formation during downstream ligation steps, and enable DNA fragments to be ligated to adaptors with complementary dT-overhangs. The desired adaptor ligated DNA size for Illumina, SOLiD and Ion Torrent platforms can be selected via gel electrophoresis before amplification by the polymerase chain reaction (PCR). New England Biolabs supplies reagents for DNA library preparation for the leading sequencing platforms. For more information, choose the DNA Library Construction Workflow tab below.
Ion Torrent™ and SOLiD™ 4 are trademarks owned by Life Technologies, Inc.
Illumina® is a registered trademark of Illumina, Inc.
454™ is a trademark of Roche.