Normalization of genomic DNA using duplex-specific nuclease

• Published in: BioTechniques
• Main Authors: Irina Shagina, Ekaterina Bogdanova, Ilgar Z. Mamedov, Yury Lebedev, Sergey Lukyanov, Dmitry Shagin
• Format: Article in Journal/Newspaper
• Language: English
• Published: Future Science Ltd 2010
• Subjects: genomic DNA; DSN normalization; duplex-specific nuclease; Biology (General); QH301-705.5; Kamchatka; Kamchatka crab
• Online Access: https://doi.org/10.2144/000113422; https://doaj.org/article/4925d55ba9154d45bb2876357e38f26a

An application of duplex-specific nuclease (DSN) normalization technology to whole-genome shotgun sequencing of genomes with a large proportion of repetitive DNA is described. The method uses a thermostable DSN from the Kamchatka crab that specifically hydrolyzes dsDNA. In model experiments on human genomic DNA, we demonstrated that DSN normalization of double-stranded DNA formed during C0t analysis is effective against abundant repetitive sequences with high sequence identity, while retaining highly divergent repeats and coding regions at base-line levels. Thus, DSN normalization applied to C0t analysis can be used to eliminate evolutionarily young repetitive elements from genomic DNA before sequencing, and should prove invaluable in studies of large eukaryotic genomes, such as those of higher plants.