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转录本与代谢物的联合鉴定:花生、野生近缘种及人工合成异源四倍体中白藜芦醇诱导与合成的见解

Coupled transcript and metabolite identification: insights on induction and synthesis of resveratrol in peanut, wild relatives and synthetic allotetraploid.

作者信息

Carvalho P A S V, Brasileiro A C, Leal-Bertioli S, Bertioli D J, Silva J P, Agostini-Costa T S, Gimenes M A

机构信息

Departamento de Genética, Instituto de Biociências, Universidade Estadual Paulista, Botucatu, SP, Brasil.

Embrapa Recursos Genéticos e Biotecnologia/CENARGEN, Parque Estação Biológica, , Brasil.

出版信息

Genet Mol Res. 2017 Sep 27;16(3):gmr-16-03-gmr.16039802. doi: 10.4238/gmr16039802.

Abstract

Resveratrol is an antioxidant that is a promising antitumoral, cardioprotective and neuroprotective agent. It has been found in a restricted number of plants including peanut (Arachis hypogaea L.) and its wild relatives. The objective of this study was to understand the relationship between resveratrol content and the expression of putative resveratrol synthase genes in four Arachis genotypes. Two diploids and two tetraploid were analyzed. Contents of resveratrol on non- and UV-treated leaves were estimated using HPLC. Resveratrol synthase (RS) was analyzed using RT-qPCR with primers developed in this study. Sequences of six Arachis species were amplified using two degenerated primer pairs that were designed based on Arachis and general RS available at GenBank. Those sequences were used to qPCR primers design. Test and control leaves were collected from plants cultivated in greenhouse and three biological replicates were evaluated for each genotype. The synthesis of resveratrol in leaves was induced by treatment with UV for 2.5 h. All genotypes studied synthesized resveratrol. Concentrations ranged from 193.66 µg/g in synthetic allotetraploid to 371.97 µg/g in A. duranensis. Natural and induced allotetraoploids showed lower levels of resveratrol than their diploid parents. Untreated samples did not produce significant amounts of resveratrol. The analysis of resveratrol content and levels of RS mRNA allowed the identification of one gene induced by the UV treatment. The data showed different amounts of RS in the different genotypes suggesting early and late response to the UV induction in the different species. The understanding of the variation found among species will help to identify species that have high resveratrol content and their ideal pos-induction times. This also will allow analysis of other tissues where high levels resveratrol would be very important, such as in seeds.

摘要

白藜芦醇是一种抗氧化剂,是一种很有前景的抗肿瘤、心脏保护和神经保护剂。它存在于包括花生(Arachis hypogaea L.)及其野生近缘种在内的少数植物中。本研究的目的是了解四种花生基因型中白藜芦醇含量与假定的白藜芦醇合酶基因表达之间的关系。分析了两个二倍体和两个四倍体。使用高效液相色谱法估计未处理和紫外线处理叶片上的白藜芦醇含量。使用本研究中开发的引物通过逆转录定量聚合酶链反应(RT-qPCR)分析白藜芦醇合酶(RS)。使用基于花生属和GenBank中可用的通用RS设计的两对简并引物扩增六种花生属物种的序列。这些序列用于设计qPCR引物。从温室种植的植物中收集测试和对照叶片,对每个基因型评估三个生物学重复。用紫外线处理2.5小时诱导叶片中白藜芦醇的合成。所有研究的基因型都合成了白藜芦醇。浓度范围从合成异源四倍体中的193.66μg/g到杜兰花生中的371.97μg/g。天然和诱导的异源四倍体的白藜芦醇水平低于其二倍体亲本。未处理的样品未产生大量白藜芦醇。对白藜芦醇含量和RS mRNA水平的分析使得能够鉴定出一个由紫外线处理诱导产生的基因。数据显示不同基因型中RS的量不同,表明不同物种对紫外线诱导的早期和晚期反应。了解物种间发现的变异将有助于识别白藜芦醇含量高的物种及其理想的诱导后时间。这也将允许分析其他白藜芦醇水平非常重要的组织,例如种子。

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