Whole genome amplification and microsatellite genotyping of herbarium DNA revealed the identity of an ancient grapevine cultivar

The Science of Nature (Impact Factor: 2.1). 09/2011; 98(9):763-772. DOI: 10.1007/s00114-011-0826-8


Reconstruction of the grapevine cultivation history has advanced tremendously during the last decade. Identification of grapevine
cultivars by using microsatellite DNA markers has mostly become a routine. The parentage of several renowned grapevine cultivars,
like Cabernet Sauvignon and Chardonnay, has been elucidated. However, the assembly of a complete grapevine genealogy is not
yet possible because missing links might no longer be in cultivation or are even extinct. This problem could be overcome by
analyzing ancient DNA from grapevine herbarium specimens and other historical remnants of once cultivated varieties. Here,
we present the first successful genotyping of a grapevine herbarium specimen and the identification of the corresponding grapevine
cultivar. Using a set of nine grapevine microsatellite markers, in combination with a whole genome amplification procedure,
we found the 90-year-old Tribidrag herbarium specimen to display the same microsatellite profile as the popular American cultivar Zinfandel. This work, together
with information from several historical documents, provides a new clue of Zinfandel cultivation in Croatia as early as the
beginning of fifteenth century, under the native name Tribidrag. Moreover, it emphasizes substantial information potential of existing grapevine and other herbarium collections worldwide.

Vitis vinifera

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    • "However, as such markers occur in non-coding regions, they are broadly unsuitable for the analysis of phenotypic traits. Microsatellite markers (also termed simple sequence repeats [SSRs]) have also been applied to aDNA samples, with recent examples including investigation of intraspecific population structure (e.g., Leino & Hagenblad, 2009) and the tracing of cultivar origin (e.g., Malenica & al., 2011). SSRs are generally more polymorphic than ITS markers. "
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    ABSTRACT: The availability of crop specimens archived in herbaria and old seed collections represent valuable resources for the analysis of plant genetic diversity and crop domestication. The ability to extract ancient DNA (aDNA) from such samples has recently allowed molecular genetic investigations to be undertaken in ancient materials. While analyses of aDNA initially focused on the use of markers which occur in multiple copies such as the internal transcribed spacer region (ITS) within ribosomal DNA and those requiring amplification of short DNA regions of variable length such as simple sequence repeats (SSRs), emphasis is now moving towards the genotyping of single nucleotide polymorphisms (SNPs), traditionally undertaken in aDNA by Sanger sequencing. Here, using a panel of barley aDNA samples previously surveyed by Sanger sequencing for putative causative SNPs within the flowering-time gene PPD-H1, we assess the utility of the Kompetitive Allele Specific PCR (KASP) genotyping platform for aDNA analysis. We find KASP to out-perform Sanger sequencing in the genotyping of aDNA samples (78% versus 61% success, respectively), as well as being robust to contamination. The small template size (≥46 bp) and one-step, closed-tube amplification/genotyping process make this platform ideally suited to the genotypic analysis of aDNA, a process which is often hampered by template DNA degradation and sample cross-contamination. Such attributes, as well as its flexibility of use and relatively low cost, make KASP particularly relevant to the genetic analysis of aDNA samples. Furthermore, KASP provides a common platform for the genotyping and analysis of corresponding SNPs in ancient, landrace and modern plant materials. The extended haplotype analysis of PPD-H1 undertaken here (allelic variation at which is thought to be important for the spread of domestication and local adaptation) provides further resolution to the previously identified geographic cline of flowering-time allele distribution, illustrating how KASP can be used to aid genetic analyses of aDNA from plant species. We further demonstrate the utility of KASP by genotyping ten additional genetic markers diagnostic for morphological traits in barley, shedding light on the phenotypic traits, alleles and allele combinations present in these unviable ancient specimens, as well as their geographic distributions.
    Taxon 08/2013; 62(4):779-789. DOI:10.12705/624.9 · 3.30 Impact Factor
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    ABSTRACT: With the expansion of molecular techniques, the historical collections have become widely used. Studying plant DNA using modern molecular techniques such as DNA sequencing plays an important role in understanding evolutionary relationships, identification through DNA barcoding, conservation status, and many other aspects of plant biology. Enormous herbarium collections are an important source of material especially for specimens from areas difficult to access or from taxa that are now extinct. The ability to utilize these specimens greatly enhances the research. However, the process of extracting DNA from herbarium specimens is often fraught with difficulty related to such variables as plant chemistry, drying method of the specimen, and chemical treatment of the specimen. Although many methods have been developed for extraction of DNA from herbarium specimens, the most frequently used are modified CTAB and DNeasy Plant Mini Kit protocols. Nine selected protocols in this chapter have been successfully used for high-quality DNA extraction from different kinds of plant herbarium tissues. These methods differ primarily with respect to their requirements for input material (from algae to vascular plants), type of the plant tissue (leaves with incrustations, sclerenchyma strands, mucilaginous tissues, needles, seeds), and further possible applications (PCR-based methods or microsatellites, AFLP).
    Methods in molecular biology (Clifton, N.J.) 01/2014; 1115:69-84. DOI:10.1007/978-1-62703-767-9_4 · 1.29 Impact Factor
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    ABSTRACT: The polyploid species complex of Rosa villosa sensu lato (Rosa section Caninae subsection Vestitae) consists of three morphologically similar polyploid species: R. sherardii, R. mollis and R. villosa. Whereas R. sherardii is distributed in central Europe, R. mollis and R. villosa represent a vicariant species pair occurring in northern Europe and in mountains of central and eastern Europe, respectively. In this study we analysed multiple data sets (morphology, cytology, microsatellites, AFLP and plastid DNA sequences) to re-evaluate the systematics of these species and to examine whether cytological differences are reflected in the taxonomy and geographical distributions. Furthermore, these data sets were used to evaluate hypotheses explaining the vicariant distribution of R. mollis and R. villosa. None of these data sets revealed a clear-cut differentiation between the species. Cytological and molecular data argued for a discrete taxonomic position of the predominantly pentaploid R. sherardii, but these data did not support a separation between the mostly tetraploid R. mollis and R. villosa. Population genetics revealed that samples of the latter species were assembled according to ploidy, but not to species affiliation or geographical distribution. Thus, we assume that the cytologically polymorphic original species had a continuous range prior to the last glaciation period and survived on nunataks or in non-glaciated coastal regions in northern Europe, but that it failed to recover its former range after the retreat of the ice sheets. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, ●●, ●●–●●.
    Botanical Journal of the Linnean Society 02/2014; 174(2). DOI:10.1111/boj.12124 · 2.53 Impact Factor
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