Strategies for Sequencing and Assembling Grapevine Genomes
Though grape transcriptomics has expanded dramatically over the last ten years, few additional novel genomic resources were developed since the release of the PN40024 reference genome in 2007. This is partly because of the difficulty associated with assembling grape genomes. Despite a relatively small genome size of ~500 Mb and modest repeat content, high sequence and structural heterozygosity makes assembling grape genomes particularly challenging. Without assemblies representative of the genetic diversity within the cultivated germplasm, identifying cultivar-specific functions not represented in the PN40024 genome has remained elusive. Now, third-generation sequencing technologies and long-range scaffolding methods have made it possible to relatively inexpensively and rapidly generate highly contiguous and complete grape genomes. This chapter will describe the challenges associated with the isolation of high-quality nucleic acids suitable for long-read sequencing and provide an overview of the sequencing and assembling approaches that can be used to successfully reconstruct grape genomes.
KeywordsGenome assembling Sequencing technologies Grapevine nucleic acid isolation
This work was supported by J. Lohr Vineyards and Wines, E. & J. Gallo Winery, Dolce Winery, the Louis P. Martini Endowment in Viticulture, and the NSF Grant #1741627. Part of this work was carried out in collaboration with UC Davis Chile and funded by the Chilean Economic Development Agency (CORFO).
- Ausubel FM, Brent R, Kingston et al (1994) Current protocols in molecular biology. Wiley, New York, pp 2.0.1–2.14.8Google Scholar
- Cheng SH, Moore BD, Seemann JR (2000) Purification of uncontaminated, intact plant RNA. In: Rapley R (ed) The nucleic acid protocols handbook. Humana Press, TotowaGoogle Scholar
- Doyle JJ, Doyle JL (1987) A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochem Bull 19:11–15Google Scholar
- Heptinstall J, Rapley R (2000) Spectrophotometric analysis of nucleic acids. In: Rapley R (ed) The Nucleic acid protocols handbook. Humana Press, TotowaGoogle Scholar
- Knebelsberger T, Stöger I (2012) DNA extraction, preservation, and amplification. In: Kress W, Erickson D (eds) DNA Barcodes. Methods in molecular biology (methods and protocols). Humana Press, TotowaGoogle Scholar
- Marsal G, Baiges I, Canals JM, Zamora F, Fort F (2013) Comparison of the efficiency of some of the most usual DNA extraction methods for woody plants in different tissues of Vitis vinifera L. J Int Sci Vigne Vin 47:227–237Google Scholar
- Minio A, Massonnet M, Figueroa-Balderas R et al (2019a) Iso-seq allows genome-independent transcriptome profiling of grape berry development. G3 Genes Genomes Genet 9:755–767Google Scholar
- Minio A, Massonnet M, Figueroa-Balderas R et al (2019b) Diploid genome assembly of the wine grape carmenere. G3 Genes Genomes Genet 9:1331–1337Google Scholar
- Ribeiro RA, Lovato MB (2007) Comparative analysis of different DNA extraction protocols in fresh and herbarium specimens of the genus Dalbergia. Gen Mol Res 6:173–187Google Scholar
- Romieu C (2010) RNA extraction from young, acidic berries and other organs from Vitis vinifera L. In: Delrot S, Medrano H, Or E, Bavaresco L, Grando S (eds) Methodologies and results in grapevine research. Springer, DordrechtGoogle Scholar
- Ruan J, Li H (2019) Fast and accurate long-read assembly with wtdbg2. BioRxiv, 530972. https://doi.org/10.1101/530972
- Zhou Y, Minio A, Massonnet M, et al (2018) Structural variants, clonal propagation, and genome evolution in grapevine (Vitis vinifera). bioRxiv 508119; doi: https://doi.org/10.1101/508119