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Article Dans Une Revue Biotechniques Année : 2016

Extraction of high-molecular-weight genomic DNA for long-read sequencing of single molecules

Résumé

De novo sequencing of complex genomes is one of the main challenges for researchers seeking high-quality reference sequences. Many de novo assemblies are based on short reads, producing fragmented genome sequences. Third-generation sequencing, with read lengths >10 kb, will improve the assembly of complex genomes, but these techniques require high-molecular-weight genomic DNA (gDNA), and gDNA extraction protocols used for obtaining smaller fragments for short-read sequencing are not suitable for this purpose. Methods of preparing gDNA for bacterial artificial chromosome (BAC) libraries could be adapted, but these approaches are time-consuming, and commercial kits for these methods are expensive. Here, we present a protocol for rapid, inexpensive extraction of high-molecular-weight gDNA from bacteria, plants, and animals. Our technique was validated using sunflower leaf samples, producing a mean read length of 12.6 kb and a maximum read length of 80 kb.

Dates et versions

hal-01608743 , version 1 (03-10-2017)

Identifiants

Citer

Baptiste Mayjonade, Jerome Gouzy, Cécile Donnadieu, Nicolas Pouilly, William Marande, et al.. Extraction of high-molecular-weight genomic DNA for long-read sequencing of single molecules. Biotechniques, 2016, 61 (4), pp.203-205. ⟨10.2144/000114460⟩. ⟨hal-01608743⟩
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