DNA Preparation In Vitis vinifera L. For Third Generation Sequencing

Abstract

BackgroundThere have been several attempts to sequence the genome of Vitis vinifera L. (grapevine), utilizing low-resolution second-generation platforms. Nevertheless, the characterization of the grapevine genetic resources and its adaptation to vulnerable conditions could be better addressed through extensive and high-resolution genome sequencing. MinION is a third-generation sequencer preferred by many laboratories due to its relatively low cost, ease of use and small size. Even though this long-read technology has been rapidly improving, to reach its full potential requires high-quality DNA. ResultsHere we establish a workflow for DNA extraction suitable for MinION sequencing long reads from grapevine. This protocol was tested with leaf samples from different positions on annual growing branches of grapevine, Purified nuclei from fresh young leaves that led to high quality, long DNA fragments, suitable for long-read sequencing were successfully generated. It is evident that longer reads in grapevine associate with both fresh tissue and adjusted conditions used for nuclei purification. ConclusionsWe propose that this workflow presents a significant advancement for long-read quality DNA isolation for grapevine and likely other plant species.