The Tubulin-Based-Polymorphism Method Provides a Simple and Effective Alternative to the Genomic Profiling of Grape

Abstract

The TBP (Tubulin-Based-Polymorphism) method, based on a nuclear ILP (Intron-Length-Polymorphism) molecular marker, has been used for genotyping 37 accessions of the genus Vitis inclusive of different species, rootstocks, wild and cultivated subspecies. A distinct DNA barcode made up by a different number of amplicons, was attributed to each of the different accessions. TBP data were compared with those obtained, with the use of an internationally validated set of six SSR markers. Genetic relationships among the different accessions, dendrogram distributions, correlation values and polymorphic index values (PICs) were definitively comparable when not in favor of TBP. Such an experimental consistency is based upon a genomic organization of the multiple members of the β-tubulin gene family, the targets of TBP-mediated amplification, that is conserved in Vitis as in any other plant species. The TBP amplicons can actually be used as a useful source of sequence polymorphisms for generating primer pairs capable of identifying specific cultivars in a simple assay. An example for the identification of the ‘Sangiovese’ cv. is reported. More generally, these data are discussed in terms of the actual advantages that the introduction of the TBP method in the field of grape characterization and genotyping can provide.

Product authenticity versus globalisation—The Tulsi case

“…Although this assay can contribute to improved transparency, because it highlights differences which align to vernacular designations of commercial products, further progress in discriminative resolution would be desirable. Given the impact of genome remodelling during the evolution of Ocimum [ 33 ], duplications in gene families would be one of the options–a principle exploited in Tubulin-Based Polymorphism, a strategy with high resolutive power [ 43 ]. For instance, the practice to commercialise Tulsi in complex mixtures with other Ayurvedic plants clearly illustrates the limitation even of trnH-psbA as trait-independent marker.…”

Intron length polymorphism of β‐tubulin genes of Aegilops biuncialis Vis

“…At the same time, TBP method saves time of the researchers and chemicals, requiring only one pair of primers, independently from the investigated taxonomic unit. Due to its simplicity TBP does not require any complicated computing, but allows to obtain a simple database of the investigated genotypes (Gavazzi et al, ). Finally, using only one pair of degenerate primers for any plant species makes TBP marker more universal than COS markers.…”

“…Variable introns are surrounded by conservative exon sites. The assay based on a simple Exon-Primed Intron-Crossing (EPIC-PCR) reaction that uses one single primer combination for any plant DNA is fast, convenient, reliable, reproducible, providing a ready-to use, and clearly intelligible results (Bardini et al, 2004;Wang et al, 2005;Braglia et al, 2010;Pirko, 2011;Gavazzi et al, 2016).…”

“…Some species of Brassica L., Lotus L., Coffea L., Eleusine Gaertn., Rosa L., etc. genera were analyzed using this system (Bardini et al, 2004;Breviario et al, 2007;Braglia et al, 2010;Galasso et al, 2010;Pirko, 2011;Gavazzi et al, 2016). At the same time, data on TBP use for the evaluation of the genetic diversity of monocotyledonous plants, particularly cereals, is incomplete.…”

A Simplified Approach for Olive (<i>Olea europaea L.</i>) Genotyping and Cultivars Traceability

“…TBP is highly successful in the molecular authentication and genomic profiling of any plant species to which a distinct Tubulin-based-Barcode (TbB) can be assigned. Depending on the way of crossing and propagation, TBP may also be very effective in the identification of varieties and hybrids as it has been already reported for grape and passionflower [10] [11]. Given the genetic history of olive and its way of propagation that brought to our times a substantially retained and ancient germplasm, it was worthwhile to verify if TBP could possibly provide a new, alternative, easy and reliable tool for the characterization of the existing genomes variability.…”