This work benefited from the networking activities coordinated within the EU-funded COST ACTION FA1106 "An integrated systems approach to determine the developmental mechanisms controlling fleshy fruit quality in tomato and grapevine". This work was supported by the 'Completamento del Centro di Genomica Funzionale Vegetale' project funded by the CARIVERONA Bank Foundation and by the 'Valorizzazione dei Principali Vitigni Autoctoni Italiani e dei loro Terroir (Vigneto)' project funded by the Italian Ministry of Agricultural and Forestry Policies. SDS was financed by the Italian Ministry of University and Research FIRB RBFR13GHC5 project "The Epigenomic Plasticity of Grapevine in Genotype per Environment Interactions".

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The authors have nothing to disclose.

本文介绍了用于解释葡萄果实风土理念代谢组学，转录和统计分析的协议。通过HPLC-ESI-MS代谢分析是不够敏感，同时检测大量的代谢产物，但相对定量是由基体效应和离子抑制/增强的影响。然而，类似的做法已经被用来形容科维纳浆果的成熟和收获后枯萎，和基体效应的修正对结果5的影响有限。此外，最近的大型多仪器实验室间研究，以确定NMR和LC-MS的用于使用相同样品非靶向代谢物组?...

根据植物的基因组，转录组，蛋白质组和代谢组大规模数据分析提供了前所未有的洞察复杂系统，如葡萄酒的风土特性反映葡萄植物及其环境之间的相互作用的行为。因为即使在相同的葡萄无性系在不同的葡萄园种植葡萄酒的风土可以是不同的，基因组学分析是没有多大用处，因为克隆的基因组是相同的。相反，它是必要看看基因表达和浆果，决定葡萄酒的品质性状的代谢特性之间的相关性。基因表达在所有转录物的化学性质相似，它通过在芯片开发通用的特点，如杂交固定的探针有利于定量分析转录福利水平的分析。与此相反，普遍在蛋白质组学分析方法第二代谢是由于个别蛋白质和代谢物的巨大的物理和化学多样性更具挑战性。在代谢组学的情况下，这种多样性是更为极端，因为个体代谢物的大小，极性，丰度和波动性大大不同，所以不存在单一的提取工艺或分析方法提供了一个全面的方法。 之间合适的非易失性代谢物的分析平台，那些基于高效液相色谱耦合质谱（HPLC-MS）比等替代性HPLC用紫外线或二极管阵列检测器（HPLC-UV，HPLC-DAD敏感得多）或核磁共振（NMR）谱，但是通过HPLC-MS定量分析可以通过的现象，如基质效应和离子抑制/增强1-3的影响。这种影响通过HPLC-MS的科维纳葡萄果实的分析过程中使用电喷雾离子源（HPLC-E的调查SI-MS），表明糖和其他分子具有最低保留时间强烈低估，也可能反映在这一区域中的大数目的分子，而其它分子的丰度可能被低估，由基体效应高估或未受影响的的，但对基体效应的数据归一化似乎对总体结果4,5-影响有限。本文所描述的方法是成熟期间积聚在高水平葡萄浆果中等极性代谢物的分析优化，这是由风土显著影响。它们包括花青素，黄酮醇，黄烷-3-醇，花青素，类黄酮等，白藜芦醇，二苯乙烯，羟基酸和羟基苯甲酸，它们共同决定了颜色，口味和葡萄酒的健康相关的属性。其它代谢物，如糖和脂族有机酸，是由于基质短跑运动员忽略，因为定量通过HPLC-MS是不可靠T和离子抑制现象5。内通过该方法选择的极性范围，该方法是在它的目的是检测许多不同的代谢物尽可能6未定位。 转录方法，使成千上万的小道消息成绩单可以同时监控由完整的葡萄基因组序列7,8的可用性提供便利。基于高通量测序的cDNA转录早期方法已经发展下一代测序的到来进入统称为RNA测序技术的过程的集合。这种方法正迅速成为首选转录研究方法。然而，基于芯片，这也让成千上万的成绩单并行通过杂交来量化大量文献，积累了葡萄。事实上，在此之前RNA测序成为一种主流技术，很多专门的商用微阵列平台已经开发允许小道消息转录，并详细检查。在广阔的各种平台，只有两个允许的全基因组转录组分析9。最进化阵列允许在单个设备上多达12个独立样品的杂交，从而减少每个实验的成本。 12个子阵列的每个组成代表29549小道消息成绩单135000 60-mer的探针。该装置已在大量的研究10-24被使用。现在这两个平台已经停产，但一个新的定制芯片最近已经设计并表示最近的发展，因为它包含代表附加新发现的基因葡萄树25探头的更大的数字。 通过转录组和代谢组学分析产生的大数据集销售需要对数据进行分析合适的统计方法，包括多元技术来确定不同形式之间的相关性s的数据的。最广泛使用的多元技术是那些基于投影，并且这些可以是无监督，如主成分分析（PCA），或监督，如双向正交投影到潜结构判别分析（O2PLS-DA）26。在这篇文章中介绍的协议采用PCA进行探索性数据分析和O2PLS-DA来确定样本的群体之间的差异。

<table><tbody><tr><td>Mill Grinder</td><td>IKA</td><td>IKA A11 basic</td><td></td></tr><tr><td>HPLC Autosampler</td><td>Beckman Coulter</td><td>&amp;nbsp;-</td><td>System Gold 508 Autosampler</td></tr><tr><td>HPLC System</td><td>Beckman Coulter</td><td>&amp;nbsp;-</td><td>System Gold 127 Solvent Module HPLC</td></tr><tr><td>C18 Guard Column</td><td>Grace</td><td>&amp;nbsp;-</td><td>Alltima HP C18 (7.5 mm x 2.1 mm; 5 &amp;mu;m) Guard Column</td></tr><tr><td>C18 Column</td><td>Grace</td><td>&amp;nbsp;-</td><td>Alltima HP C18 (150 mm x 2.1 mm; 3 &amp;mu;m) Column</td></tr><tr><td>Mass Spectometer</td><td>Bruker Daltonics</td><td>&amp;nbsp;-</td><td>Bruker Esquire 6000; The mass spectometer was equipped with an ESI source and the analyzer was an ion trap.</td></tr><tr><td>Extraction solvents and HPLC buffers</td><td>Sigma</td><td>34966</td><td>Methanol LC-MS grade</td></tr><tr><td>Sigma</td><td>94318</td><td>Formic acid LC-MS grade</td><td></td></tr><tr><td>Sigma</td><td>34967</td><td>Acetonitrile LC-MS grade</td><td></td></tr><tr><td>Sigma</td><td>39253</td><td>Water&amp;nbsp; LC-MS grade</td><td></td></tr><tr><td>Minisart RC 4 Syringe filters (0.2 &amp;mu;m)</td><td>Sartorius</td><td>17764</td><td></td></tr><tr><td>Softwares for data collection (a) and processing (b)</td><td>Bruker Daltonics</td><td>-</td><td>Bruker Daltonics Esquire 5.2 Control (a); Esquire 3.2 Data Analysis and MzMine 2.2 softwares (b)</td></tr><tr><td>Spectrum Plant Total RNA kit</td><td>Sigma-Aldrich</td><td>STRN250-1KT</td><td>For total RNA extractino from grape pericarps</td></tr><tr><td>Nanodrop 1000</td><td>Thermo Scientific</td><td>1000</td><td></td></tr><tr><td>BioAnalyzer 2100</td><td>Agilent Technologies</td><td>G2939A</td><td></td></tr><tr><td>RNA 6000 Nano Reagents</td><td>Agilent Technologies</td><td>5067-1511</td><td></td></tr><tr><td>RNA Chips</td><td>Agilent Technologies</td><td>5067-1511</td><td></td></tr><tr><td>Agilent Gene Expression Wash Buffer 1</td><td>Agilent Technologies</td><td>5188-5325</td><td></td></tr><tr><td>Agilent Gene Expression Wash Buffer 2</td><td>Agilent Technologies</td><td>5188-5326</td><td></td></tr><tr><td>LowInput QuickAmp Labeling kit One-Color</td><td>Agilent Technologies</td><td>5190-2305</td><td></td></tr><tr><td>Kit RNA Spike In - One-Color</td><td>Agilent Technologies</td><td>5188-5282</td><td></td></tr><tr><td>Gene Expression Hybridization Kit</td><td>Agilent Technologies</td><td>5188-5242</td><td></td></tr><tr><td>RNeasy Mini Kit (50)</td><td>Qiagen</td><td>74104</td><td>For cRNA Purification</td></tr><tr><td>Agilent SurePrint HD 4X44K 60-mer Microarray</td><td>Agilent Technologies</td><td>G2514F-048771&amp;nbsp;</td><td></td></tr><tr><td>eArray</td><td>Agilent Technologies</td><td>-</td><td>&lt;a href=&quot;https://earray.chem.agilent.com/earray/&quot;&gt;https://earray.chem.agilent.com/earray/&lt;/a&gt;</td></tr><tr><td>Gasket slides</td><td>Agilent Technologies</td><td>G2534-60012</td><td>Enable Agilent SurePrint Microarray 4-array Hybridization</td></tr><tr><td>Thermostatic bath</td><td>Julabo</td><td>-</td><td></td></tr><tr><td>Hybridization Chamber</td><td>Agilent Technologies</td><td>G2534-60001</td><td></td></tr><tr><td>Microarray Hybridization Oven</td><td>Agilent Technologies</td><td>G2545A</td><td></td></tr><tr><td>Hybridization Oven Rotator Rack</td><td>Agilent Technologies</td><td>G2530-60029</td><td></td></tr><tr><td>Rotator Rack Conversion Rod</td><td>Agilent Technologies</td><td>G2530-60030</td><td></td></tr><tr><td>Staining kit</td><td>Bio-Optica</td><td>10-2000</td><td>Slide-staining dish and Slide rack</td></tr><tr><td>Magnetic stirrer device</td><td>AREX Heating Magnetic Stirrer</td><td>F20540163&amp;nbsp;</td><td></td></tr><tr><td>Thermostatic Oven</td><td>Thermo Scientific</td><td>Heraeus - 6030</td><td></td></tr><tr><td>Agilent Microarray Scanner</td><td>Agilent Technologies</td><td>G2565CA</td><td></td></tr><tr><td>Scanner Carousel, 48-position</td><td>Agilent Technologies</td><td>G2505-60502</td><td></td></tr><tr><td>Slide Holders</td><td>Agilent Technologies</td><td>G2505-60525</td><td></td></tr><tr><td>Feature extraction software v11.5</td><td>Agilent Technologies</td><td>-</td><td>inside the Agilent Microarray Scanner G2565CA</td></tr><tr><td>SIMCA + V13 Software</td><td>Umetrics</td><td></td><td></td></tr></tbody></table>

the terroir concept interpreted through grape berry metabolomics and transcriptomics

风土指的是，根据特定的生境和管理实践影响作物的特性，如葡萄（ 葡萄 ）的环境因素的结合。本文将介绍如何某些风土签名可以使用多元统计分析小道消息科维纳品种的浆果代谢和转录组进行检测。该方法首先需要一个适当的采样计划。在这个案例研究中，科维纳品种的特定克隆选择遗传差异最小化，并从代表在三个不同的生长季节三种不同的宏观区域7葡萄园收集样品。一个不相关的LC-MS代谢物组学的方法是，由于其高灵敏度，建议使用MZmine软件和基于碎片树分析的代谢物的识别策略伴有有效的数据处理。可以使用微阵列可以实现全面的转录组分析含探针涵盖〜所有预测葡萄基因的99％，允许所有的差异表达的基因在不同风土的上下文中同时分析。最后，基于投影的方法多元数据分析可以被用来克服强特异性复古效果，允许代谢和转录数据被集成并详细分析以识别信息相关性。

本文中介绍的案例研究产生了最终的数据矩阵包括552信号（M / Z功能），包括分子离子以及它们的同位素，加合物和一些片断，其中189样本相对量化（7葡萄园×3成熟期×3生长季点¯x 3生物学重复）。因此104328用于数据点的总数为。碎片树分析导致282 M / Z功能注释，相应的代谢产物加合物，同位素和片段。由主成分分析整个数据矩阵的探索性分析表明根据熟化?...

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The Terroir Concept Interpreted through Grape Berry Metabolomics and Transcriptomics

本文描述了为了深入了解风土概念， 即非靶向代谢组学，转录和多元统计分析，以葡萄果实成绩单和代谢产物的应用中，环境对果实品质性状的影响。

风土通过概念葡萄果实代谢组学和转录组解释的

Terroir refers to the combination of environmental factors that affect the characteristics of crops such as grapevine (Vitis vinifera) according to ...

the-terroir-concept-interpreted-through-grape-berry-metabolomics

Universidad San Sebastian

Research

JoVE Journal

Genetics

10.3K 次观看.  University of Verona. 本文描述了为了深入了解风土概念， 即非靶向代谢组学，转录和多元统计分析，以葡萄果实成绩单和代谢产物的应用中，环境对果实品质性状的影响。 

视频: The Terroir Concept Interpreted through Grape Berry Metabolomics and Transcriptomics

In Vitro Transcription Assays and Their Application in Drug Discovery

In vitro transcription assays have been developed and widely used for many years to study the molecular mechanisms involved in transcription. This process requires multi-subunit DNA-dependent RNA polymerase (RNAP) and a series of transcription factors that act to modulate the activity of RNAP during gene expression. Sequencing gel electrophoresis of radiolabeled transcripts is used to provide detailed mechanistic information on how transcription proceeds and what parameters can affect it. In this paper we describe the protocol to study how the essential elongation factor NusA regulates transcriptional pausing, as well as a method to identify an antibacterial agent targeting transcription initiation through inhibition of RNAP holoenzyme formation. These methods can be used a as platform for the development of additional approaches to explore the mechanism of action of the transcription factors which still remain unclear, as well as new antibacterial agents targeting transcription which is an underutilized drug target in antibiotic research and development.

In Vitro Transcription Assays and Their Application in Drug Discovery

In this manuscript, we describe a protocol to functionally examine transcription and the inhibitory activity of antibacterial agents targeting bacterial transcription.

体外转录测定及其在药物研究中的应用

In vitro transcription assays have been developed and widely used for many years to study the molecular mechanisms involved in transcription. This process ...

科研

遗传学

Delivery of Nucleic Acids through Embryo Microinjection in the Worldwide Agricultural Pest Insect, Ceratitis capitata

The Mediterranean fruit fly (medfly) Ceratitis capitata (Wiedemann) (Diptera: Tephritidae) is a pest species with extremely high agricultural relevance. This is due to its reproductive behavior: females damage the external surface of fruits and vegetables when they lay eggs and the hatched larvae feed on their pulp. Wild C. capitata populations are traditionally controlled through insecticide spraying and/or eco-friendly approaches, the most successful being the Sterile Insect Technique (SIT). The SIT relies on mass-rearing, radiation-based sterilization and field release of males that retain their capacity to mate but are not able to generate fertile progeny. The advent and the subsequent rapid development of biotechnological tools, together with the availability of the medfly genome sequence, has greatly boosted our understanding of the biology of this species. This favored the proliferation of new strategies for genome manipulation, which can be applied to population control.
In this context, embryo microinjection plays a dual role in expanding the toolbox for medfly control. The ability to interfere with the function of genes that regulate key biological processes, indeed, expands our understanding of the molecular machinery underlying medfly invasiveness. Furthermore, the ability to achieve germ-line transformation facilitates the production of multiple transgenic strains that can be tested for future field applications in novel SIT settings. Indeed, genetic manipulation can be used to confer desirable traits that can, for example, be used to monitor sterile male performance in the field, or that can result in early life-stage lethality. Here we describe a method to microinject nucleic acids into medfly embryos to achieve these two main goals.

Delivery of Nucleic Acids through Embryo Microinjection in the Worldwide Agricultural Pest Insect, Ceratitis capitata

The Mediterranean fruit fly (medfly) Ceratitis capitata (Diptera: Tephritidae) is a worldwide pest of agriculture. A deeper understanding of its biology is key to control medfly populations and thus reduce economic impact. Embryo microinjection is a fundamental tool allowing both germ-line transformation and reverse genetics studies in this species.

核酸的传递通过显微注射胚胎在世界范围内的农业害虫，<em&gt;地中海实蝇</em

The Mediterranean fruit fly (medfly) Ceratitis capitata (Wiedemann) (Diptera: Tephritidae) is a pest species with extremely high agricultural relevance. ...

Leveraging CyVerse Resources for De Novo Comparative Transcriptomics of Underserved (Non-model) Organisms

This workflow allows novice researchers to leverage advanced computational resources such as cloud computing to carry out pairwise comparative transcriptomics. It also serves as a primer for biologists to develop data scientist computational skills, e.g. executing bash commands, visualization and management of large data sets. All command line code and further explanations of each command or step can be found on the wiki (<a href="https://wiki.cyverse.org/wiki/x/dgGtAQ">https://wiki.cyverse.org/wiki/x/dgGtAQ</a>). The Discovery Environment and Atmosphere platforms are connected together through the CyVerse Data Store. As such, once the initial raw sequencing data has been uploaded there is no more need to transfer large data files over an Internet connection, minimizing the amount of time needed to conduct analyses. This protocol is designed to analyze only two experimental treatments or conditions. Differential gene expression analysis is conducted through pairwise comparisons, and will not be suitable to test multiple factors. This workflow is also designed to be manual rather than automated. Each step must be executed and investigated by the user, yielding a better understanding of data and analytical outputs, and therefore better results for the user. Once complete, this protocol will yield de novo assembled transcriptome(s) for underserved (non-model) organisms without the need to map to previously assembled reference genomes (which are usually not available in underserved organism). These de novo transcriptomes are further used in pairwise differential gene expression analysis to investigate genes differing between two experimental conditions. Differentially expressed genes are then functionally annotated to understand the genetic response organisms have to experimental conditions. In total, the data derived from this protocol is used to test hypotheses about biological responses of underserved organisms.

Leveraging CyVerse Resources for De Novo Comparative Transcriptomics of Underserved Non-model Organisms

This protocol outlines a comparative de novo transcriptome assembly and annotation workflow for novice bioinformaticians. The workflow is available for free entirely through CyVerse and connected by the Data Store. Command line and graphical user interfaces are used, but all code needed is available to copy and paste.

利用CyVerse资源<em&gt; De Novo</em&gt;不合格（非模型）生物的比较转录组学

This workflow allows novice researchers to leverage advanced computational resources such as cloud computing to carry out pairwise comparative ...

Controllable Ion Channel Expression through Inducible Transient Transfection

Transfection, the delivery of foreign nucleic acids into a cell, is a powerful tool in protein research. Through this method, ion channels can be investigated through electrophysiological analysis, biochemical characterization, mutational studies, and their effects on cellular processes. Transient transfections offer a simple protocol in which the protein becomes available for analysis within a few hours to days. Although this method presents a relatively straightforward and time efficient protocol, one of the critical components is calibrating the expression of the gene of interest to physiological relevant levels or levels that are suitable for analysis. To this end, many different approaches that offer the ability to control the expression of the gene of interest have emerged. Several stable cell transfection protocols provide a way to permanently introduce a gene of interest into the cellular genome under the regulation of a tetracycline-controlled transcriptional activation. While this technique produces reliable expression levels, each gene of interest requires a few weeks of skilled work including calibration of a killing curve, selection of cell colonies, and overall more resources. Here we present a protocol that uses transient transfection of the Transient Receptor Potential cation channel subfamily V member 1 (TRPV1) gene in an inducible system as an efficient way to express a protein in a controlled manner which is essential in ion channel analysis. We demonstrate that using this technique, we are able to perform calcium imaging, whole cell, and single channel analysis with controlled channel levels required for each type of data collection with a single transfection. Overall, this provides a replicable technique that can be used to study ion channels structure and function.

Studying ion channels through a heterologously expressing system has become a core technique in biomedical research. In this manuscript, we present a time efficient method to achieve tightly controlled ion channel expression by performing transient transfection under the control of an inducible promoter.

通过诱导瞬时转染可控离子通道表达

Transfection, the delivery of foreign nucleic acids into a cell, is a powerful tool in protein research. Through this method, ion channels can be ...

Transcriptomic Analysis of C. elegans RNA Sequencing Data Through the Tuxedo Suite on the Galaxy Project

Next generation sequencing (NGS) technologies have revolutionized the nature of biological investigation. Of these, RNA Sequencing (RNA-Seq) has emerged as a powerful tool for gene-expression analysis and transcriptome mapping. However, handling RNA-Seq datasets requires sophisticated computational expertise and poses inherent challenges for biology researchers. This bottleneck has been mitigated by the open access Galaxy project that allows users without bioinformatics skills to analyze RNA-Seq data, and the Database for Annotation, Visualization, and Integrated Discovery (DAVID), a Gene Ontology (GO) term analysis suite that helps derive biological meaning from large data sets. However, for first-time users and bioinformatics' amateurs, self-learning and familiarization with these platforms can be time-consuming and daunting. We describe a straightforward workflow that will help C. elegans researchers to isolate worm RNA, conduct an RNA-Seq experiment and analyze the data using Galaxy and DAVID platforms. This protocol provides stepwise instructions for using the various Galaxy modules for accessing raw NGS data, quality-control checks, alignment, and differential gene expression analysis, guiding the user with parameters at every step to generate a gene list that can be screened for enrichment of gene classes or biological processes using DAVID. Overall, we anticipate that this article will provide information to C. elegans researchers undertaking RNA-Seq experiments for the first time as well as frequent users running a small number of samples.

Transcriptomic Analysis of C. elegans RNA Sequencing Data Through the Tuxedo Suite on the Galaxy Project

Galaxy and DAVID have emerged as popular tools that allow investigators without bioinformatics training to analyze and interpret RNA-Seq data. We describe a protocol for C. elegans researchers to perform RNA-Seq experiments, access and process the dataset using Galaxy and obtain meaningful biological information from the gene lists using DAVID.

的转录分析<em&gt;ç</em&gt;。<em&gt;线虫</em&gt; RNA测序数据通过炫套房的银河项目

Next generation sequencing (NGS) technologies have revolutionized the nature of biological investigation. Of these, RNA Sequencing (RNA-Seq) has emerged ...

Untargeted Liquid Chromatography-Mass Spectrometry-Based Metabolomics Analysis of Wheat Grain

Understanding the interactions between genes, the environment and management in agricultural practice could allow more accurate prediction and management of product yield and quality. Metabolomics data provides a read-out of these interactions at a given moment in time and is informative of an organism&#39;s biochemical status. Further, individual metabolites or panels of metabolites can be used as precise biomarkers for yield and quality prediction and management. The plant metabolome is predicted to contain thousands of small molecules with varied physicochemical properties that provide an opportunity for a biochemical insight into physiological traits and biomarker discovery. To exploit this, a key aim for metabolomics researchers is to capture as much of the physicochemical diversity as possible within a single analysis. Here we present a liquid chromatography-mass spectrometry-based untargeted metabolomics method for the analysis of field-grown wheat grain. The method uses the liquid chromatograph quaternary solvent manager to introduce a third mobile phase and combines a traditional reversed-phase gradient with a lipid-amenable gradient. Grain preparation, metabolite extraction, instrumental analysis and data processing workflows are described in detail. Good mass accuracy and signal reproducibility were observed, and the method yielded approximately 500 biologically relevant features per ionization mode. Further, significantly different metabolite and lipid feature signals between wheat varieties were determined.

A method for the untargeted analysis of wheat grain metabolites and lipids is presented. The protocol includes an acetonitrile metabolite extraction method and reversed phase liquid chromatography-mass spectrometry methodology, with acquisition in positive and negative electrospray ionization modes.

无目标液相色谱-质量光谱学-小麦粒基代谢代谢学分析

Understanding the interactions between genes, the environment and management in agricultural practice could allow more accurate prediction and management ...

生物化学

Profiling Volatile Compounds in Blackcurrant Fruit using Headspace Solid-Phase Microextraction Coupled to Gas Chromatography-Mass Spectrometry

There is an increasing interest in measuring volatile organic compounds (VOCs) emitted by ripe fruits for the purpose of breeding varieties or cultivars with enhanced organoleptic characteristics and thus, to increase consumer acceptance. High-throughput metabolomic platforms have been recently developed to quantify a wide range of metabolites in different plant tissues, including key compounds responsible for fruit taste and aroma quality (volatilomics). A method using headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry (GC-MS) is described here for the identification and quantification of VOCs emitted by ripe blackcurrant fruits, a berry highly appreciated for its flavor and health benefits.
Ripe fruits of blackcurrant plants (Ribes nigrum) were harvested and directly frozen in liquid nitrogen. After tissue homogenization to produce a fine powder, samples were thawed and immediately mixed with sodium chloride solution. Following centrifugation, the supernatant was transferred into a headspace glass vial containing sodium chloride. VOCs were then extracted using a solid-phase microextraction (SPME) fiber and a gas chromatograph coupled to an ion trap mass spectrometer. Volatile quantification was performed on the resulting ion chromatograms by integrating peak area, using a specific m/z ion for each VOC. Correct VOC annotation was confirmed by comparing retention times and mass spectra of pure commercial standards run under the same conditions as the samples. More than 60 VOCs were identified in ripe blackcurrant fruits grown in contrasting European locations. Among the identified VOCs, key aroma compounds, such as terpenoids and C6 volatiles, can be used as biomarkers for blackcurrant fruit quality. In addition, advantages and disadvantages of the method are discussed, including prospective improvements. Furthermore, the use of controls for batch correction and minimization of drift intensity have been emphasized.

A headspace solid-phase microextraction-gas-chromatography platform is described here for fast, reliable, and semi-automated volatile identification and quantification in ripe blackcurrant fruits. This technique can be used to increase knowledge about fruit aroma and to select cultivars with enhanced flavor for the purpose of breeding.

使用顶空固相微萃取与气相色谱-质谱联用分析黑醋栗果实中的挥发性化合物

There is an increasing interest in measuring volatile organic compounds (VOCs) emitted by ripe fruits for the purpose of breeding varieties or cultivars ...

Annotation of Plant Gene Function via Combined Genomics, Metabolomics and Informatics

Given the ever expanding number of model plant species for which complete genome sequences are available and the abundance of bio-resources such as knockout mutants, wild accessions and advanced breeding populations, there is a rising burden for gene functional annotation. In this protocol, annotation of plant gene function using combined co-expression gene analysis, metabolomics and informatics is provided (Figure 1). This approach is based on the theory of using target genes of known function to allow the identification of non-annotated genes likely to be involved in a certain metabolic process, with the identification of target compounds via metabolomics. Strategies are put forward for applying this information on populations generated by both forward and reverse genetics approaches in spite of none of these are effortless. By corollary this approach can also be used as an approach to characterise unknown peaks representing new or specific secondary metabolites in the limited tissues, plant species or stress treatment, which is currently the important trial to understanding plant metabolism.

Combination of genomics, co-expression gene analysis and the identification of target compounds via metabolism give gene functional annotation.

联合基因组学，代谢组学和信息通过植物基因功能的注解

Given the ever expanding number of model plant species for which complete genome sequences are available and the abundance of bio-resources such as ...

生物学

Comprehensive Workflow for the Genome-wide Identification and Expression Meta-analysis of the ATL E3 Ubiquitin Ligase Gene Family in Grapevine

Classification and nomenclature of genes in a family can significantly contribute to the description of the diversity of encoded proteins and to the prediction of family functions based on several features, such as the presence of sequence motifs or of particular sites for post-translational modification and the expression profile of family members in different conditions. This work describes a detailed protocol for gene family characterization. Here, the procedure is applied to the characterization of the Arabidopsis T&#243;xicos in Levadura (ATL) E3 ubiquitin ligase family in grapevine. The methods include the genome-wide identification of family members, the characterization of gene localization, structure, and duplication, the analysis of conserved protein motifs, the prediction of protein localization and phosphorylation sites as well as gene expression profiling across the family in different datasets. Such procedure, which could be extended to further analyses depending on experimental purposes, could be applied to any gene family in any plant species for which genomic data are available, and it provides valuable information to identify interesting candidates for functional studies, giving insights into the molecular mechanisms of plant adaptation to their environment.

This article describes the procedure for the identification and characterization of a gene family in grapevine applied to the family of Arabidopsis T&#243;xicos in Levadura (ATL) E3 ubiquitin ligases.

葡萄全基因组鉴定与表达的综合工作流 E3 泛素连接基因家族的 Meta 分析

Classification and nomenclature of genes in a family can significantly contribute to the description of the diversity of encoded proteins and to the ...

使用 Amplicon 宏基因组学分析葡萄中的细菌群落

Profiling the Bacterial Community of Fermenting Traminette Grapes during Wine Production using Metagenomic Amplicon Sequencing

Advances in sequencing technology and the relatively easy access to the use of bioinformatics tools to profile microbial community structures have facilitated a better understanding of both culturable and non-culturable microbes in grapes and wine. During industrial fermentation, microbes, known and unknown, are often responsible for product development and off-flavor. Therefore, profiling the bacteria from grape to wine can enable an easy understanding of in situ microbial dynamics. In this study, the bacteria of Traminette grapes must undergoing fermentation, and the final wine were subjected to DNA extraction that yielded 15 ng/&#181;L to 87 ng/&#181;L. The 16S amplicon of the hypervariable region of the V4 region was sequenced, relatively abundant bacteria consisting of phyla Proteobacteria, Actinobacteriota, Firmicutes, Bacteroidota, Fusobacteriota and followed by the Verrucomicrobiota, Halobacterota, Desulfobacterota, Myxococcota, and Acidobacteriota. A Venn diagram analysis of the shared unique operational taxonomic units (OTU) revealed that 15 bacteria phyla were common to both grape must, fermenting stage, and final wine. Phyla that were not previously reported were detected using the 16S amplicon sequencing, as well as genera such as Enterobacteriaceae and Lactobacillaceae. Variation in the organic nutrient use in wine and its impact on bacteria was tested; Traminette R tank containing Fermaid O and Traminette L stimulated with Stimula Sauvignon blanc + Fermaid O. Alpha diversity using the Kruskal-Wallis test determined the degree of evenness. The beta diversity indicated a shift in the bacteria at the fermentation stage for the two treatments, and the final wine bacteria looked similar. The study confirmed that 16S amplicon sequencing can be used to monitor bacteria changes during wine production to support quality and better utilization of grape bacteria during wine production.

作者聚焦：通过下一代测序探索发酵微生物组

Here, we present a protocol to describe amplicon metagenomic for determining the bacterial community of Traminette grapes, fermenting grapes, and final wine.

风土通过概念葡萄果实代谢组学和转录组解释的

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联合基因组学，代谢组学和信息通过植物基因功能的注解

葡萄全基因组鉴定与表达的综合工作流 E3 泛素连接基因家族的 Meta 分析

使用宏基因组扩增子测序分析葡萄酒生产过程中发酵Traminette葡萄的细菌群落