A thaumatin-like genomic sequence identification in Vitis vinifera l., stormy wines and musts based on direct pcr

Authors

  • Jana Žiarovská Slovak University of Agriculture, Faculty of Agrobiology and Food Resources, Department of Genetics and Plant Breeding, Tr. A. Hlinku 2, 949 76 Nitra
  • Lucia Zeleňáková Slovak University of Agriculture, Faculty of Biotechnology and Food Sciences, Department of Food Hygiene and Safety, Tr. A. Hlinku 2, 949 76 Nitra
  • Miroslava Kačániová Slovak University of Agriculture, Faculty of Biotechnology and Food Sciences, Department of Microbiology, Tr. A. Hlinku 2, 949 76 Nitra, Slovakia
  • Eloy Fernández Cusimamani Czech University of Life Sciences Prague, Faculty of Tropical AgriSciences, Kamýcká 961/129, 165 00, Prague-Suchdol

DOI:

https://doi.org/10.5219/892

Keywords:

direct PCR, Vitis vinifera L., thaumatin-like sequence, stormy wine, must

Abstract

Direct polymerase chain reaction method was use to amplify a thaumatin-like sequence of Vitis vinifera L. in grapes as well as in stormy wines and musts. Thaumatin-like proteins (TLPs) of Vitis vinifera possess beside its function in abiotic and biotic stress response another one - they are able to cause protein haze in wine unless removed prior to bottling. Direct PCR is an approach where omission of DNA extraction is typical prior the amplification of the target site of plant genome. Crude extract or small pieces of plant tissues are used in the analysis directly without steps of extraction and purification of gDNA. The biological material that was used in analysis was collected during August - October 2017 in local stores and winery Sabo and comprises from cultivars Iršai, Muškát, Savignon Blanc, Svätovavrinecké, Dornfelder and Pálava. Direct PCR was performed by a cutted piece of grape tissue and a dilution buffer was use in 1:2 for stormy wine or must, respectively. Direct amplification of thaumatin-like protein sequence of Vitis vinifera was performed along with the control reactions with the primers for conserved region of plant chloroplast. Possitive amplification of thaumatin-like allergen sequence resulted in 570 bp amplicon. The most abundant amplicons were amplified in stormy wines, followed by musts and the amplicons from grapes were weaker when comparing them to others. The amplicon specificity checking of obtained PCR product of thaumatin-like allergen was performed by restriction cleavage by Psi I and resulted in restriction amplicons of the 80 bp, 81 bp, 94 bp and 315 bp in length. Confirmation of the amplicon specificity by restriction cleavage support the potential of direct PCR to become a reproducible method that will be fully applicable in routine analysis of not only plant genomes in the future, but it was demonstrated, that it works in liquids, too. 

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Published

2018-03-05

How to Cite

Žiarovská, J. ., Zeleňáková, L. ., Kačániová, M. ., & Cusimamani, E. F. (2018). A thaumatin-like genomic sequence identification in Vitis vinifera l., stormy wines and musts based on direct pcr. Potravinarstvo Slovak Journal of Food Sciences, 12(1), 226–232. https://doi.org/10.5219/892

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