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替代 SNP 检测平台，HRM 和生物传感器，用于使用 F3H 和 LDOX 基因进行葡萄属品种鉴定。

Alternative SNP detection platforms, HRM and biosensors, for varietal identification in Vitis vinifera L. using F3H and LDOX genes.

机构信息

University of Trás-os-Montes and Alto Douro, P.O. Box 1013, 5000-911, Vila Real, Portugal.

University of Lisboa, Faculty of Sciences, BioISI - Biosystems & Integrative Sciences Institute, Campo Grande, Lisboa, Portugal.

出版信息

Sci Rep. 2018 Apr 11;8(1):5850. doi: 10.1038/s41598-018-24158-9.

DOI:10.1038/s41598-018-24158-9
PMID:29643453
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC5895793/
Abstract

The wine sector requires quick and reliable methods for Vitis vinifera L. varietal identification. The number of V. vinifera varieties is estimated in about 5,000 worldwide. Single Nucleotide Polymorphisms (SNPs) represent the most basic and abundant form of genetic sequence variation, being adequate for varietal discrimination. The aim of this work was to develop DNA-based assays suitable to detect SNP variation in V. vinifera, allowing varietal discrimination. Genotyping by sequencing allowed the detection of eleven SNPs on two genes of the anthocyanin pathway, the flavanone 3-hydroxylase (F3H, EC: 1.14.11.9), and the leucoanthocyanidin dioxygenase (LDOX, EC 1.14.11.19; synonym anthocyanidin synthase, ANS) in twenty V. vinifera varieties. Three High Resolution Melting (HRM) assays were designed based on the sequencing information, discriminating five of the 20 varieties: Alicante Bouschet, Donzelinho Tinto, Merlot, Moscatel Galego and Tinta Roriz. Sanger sequencing of the HRM assay products confirmed the HRM profiles. Three probes, with different lengths and sequences, were used as bio-recognition elements in an optical biosensor platform based on a long period grating (LPG) fiber optic sensor. The label free platform detected a difference of a single SNP using genomic DNA samples. The two different platforms were successfully applied for grapevine varietal identification.

摘要

葡萄酒行业需要快速可靠的方法来鉴定 Vitis vinifera L. 品种。全球估计有 5000 多种 V. vinifera 品种。单核苷酸多态性 (SNP) 是遗传序列变异的最基本和最丰富的形式，足以进行品种鉴别。本工作旨在开发基于 DNA 的检测方法，以检测 V. vinifera 中的 SNP 变异，从而实现品种鉴别。测序基因分型允许在两个花青素途径基因上检测到 11 个 SNP，即类黄酮 3-羟化酶 (F3H，EC:1.14.11.9) 和无色花青素双加氧酶 (LDOX，EC 1.14.11.19；同义词：花青素合酶，ANS)，在 20 个 V. vinifera 品种中。基于测序信息设计了三个高分辨率熔解 (HRM) 分析，可区分 20 个品种中的 5 个品种： Alicante Bouschet、Donzelinho Tinto、Merlot、Moscatel Galego 和 Tinta Roriz。HRM 分析产物的 Sanger 测序证实了 HRM 图谱。三种不同长度和序列的探针用作基于长周期光栅 (LPG) 光纤传感器的光学生物传感器平台的生物识别元件。无标记平台使用基因组 DNA 样本检测到单个 SNP 的差异。两种不同的平台都成功应用于葡萄品种鉴定。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/8d4734aacdb0/41598_2018_24158_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/6bfb6c02975a/41598_2018_24158_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/8832d275b50d/41598_2018_24158_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/08d592bb4c38/41598_2018_24158_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/aea40c95285d/41598_2018_24158_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/a48894eb7991/41598_2018_24158_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/8d4734aacdb0/41598_2018_24158_Fig6_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/6bfb6c02975a/41598_2018_24158_Fig1_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/8832d275b50d/41598_2018_24158_Fig2_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/08d592bb4c38/41598_2018_24158_Fig3_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/aea40c95285d/41598_2018_24158_Fig4_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/a48894eb7991/41598_2018_24158_Fig5_HTML.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e67/5895793/8d4734aacdb0/41598_2018_24158_Fig6_HTML.jpg

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