An efficient and rapid protocol for plant nuclear DNA preparation suitable for next generation sequencing methods
2011
Carrier, Gregory | Santoni, Sylvain | Rodier-Goud, Marguerite | Canaguier, Aurélie | Kochko, Alexandre de | Dubreuil-Tranchant, Christine | This, Patrice | Boursiquot, Jean-Michel | Cunff, Loïc Le
Premise of the study: In this study, we developed a nuclear DNA extraction protocol for Next Generation Sequencers (NGS). METHODS: and Results: We applied this extraction method to grapevines and coffee trees, which are known to contain many secondary metabolites. The nuclear DNA obtained was sequenced by the 454/GS-FLX method. We obtained excellent results, with less than 4% cytoplasmic DNA, in a similar way to a BAC (Bacterial Artificial Chromosome)-building protocol. We also compared our protocol with a classic DNA extraction using specific cytoplasmic DNA amplification. Results showed a lower cytoplasmic DNA contamination with the new protocol. CONCLUSIONS: The method presented here is fast and economical. The DNA obtained is of high quality, with a low level of cytoplasmic DNA contamination, and very efficient for the construction of sequencing libraries.
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