Plant Proteomics Methods and Protocols
Plant Proteomics Methods and Protocols
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DOI：
10.1007/978-1-62703-631-3_1
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发表时间：
2014-01-01
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影响因子：
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通讯作者：
Jorrin-Novo, Jesus V.
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作者：
Jorrin-Novo, Jesus V.
在第一章，引言中，它打算从方法论的角度总结植物蛋白质组学的最新进展，重点是基于质谱学的策略。因此，本章主要针对初学者或那些试图进入该领域的人，而不是那些在植物蛋白质组学研究方面具有实际经验或长期发展轨迹的人。从实验设计到数据分析的不同替代工作流程、方法、技术和方案将被简要评论，并交叉引用以前的专著和评论，以及本书的其余章节。还将简要讨论处理蛋白质的困难，以及这种方法的威力、局限性和挑战。蛋白质作为分子实体，以及细胞蛋白质组作为一个整体，比我们过去认为的要复杂得多，可以在一个实验中进行研究。正因为如此，对它的研究需要采取分级和互补的策略。对复杂样品的MS分析可能会产生多达100,000个用标准程序无法轻松分析的多肽光谱。因此，蛋白质组学比其他组学更需要枯燥的实验室、时间和数据挖掘的努力。作为主要结论，可以说蛋白质组学正处于起步阶段。它开始为正确的基因注释、鉴定和鉴定基因产品或蛋白质物种以及了解活的生物体做出重要贡献，并在翻译研究中具有巨大的应用潜力。然而，尽管它有巨大的潜力，就像在任何其他实验方法中一样，它远不是潘多拉的盒子。就植物研究而言，蛋白质组学的全部潜力还远未完全被开发，第二代、第三代和第四代蛋白质组学技术的使用仍然非常有限。到目前为止发表的大多数植物蛋白质组学论文属于描述性、亚细胞性和比较蛋白质组学亚组，主要使用少数实验模型系统-那些其基因组已被测序的系统-从生物学的角度来看，具有很强的描述性和推测性。从现在起，我们应该更加重视翻译后蛋白质组学和相互作用组学的研究，并转向有针对性的、假设驱动的方法。此外，更重要的是，我们应该通过其他组学或经典的生化策略进行数据验证，试图对细胞生物学有更深、更真实、更准确的看法和理解。在现代系统生物学的概念中，蛋白质组学必须被视为全球多学科方法的一部分。要使蛋白质组学实验具有生物学意义，需要一个适当的实验设计、数据验证、解释和发表政策。
In this first, introductory chapter, it is intended to summarize from a methodological point of view the state of the art in plant proteomics, focusing on mass spectrometry-based strategies. Thus, this chapter is mainly directed at beginners or at those trying to get into the field, rather than at those with real experience or a long trajectory in plant proteomics research. The different alternative workflows, methods, techniques, and protocols from the experimental design to the data analysis will be briefly commented, with cross references to previous monographs and reviews, as well as to the rest of the book chapters. The difficulty of working with proteins, together with the power, limitations, and challenges of the approach will also be briefly discussed.Proteins, as molecular entities, and the cell proteome, as a whole, are much more complex than what we thought in the past and can be studied in a single experiment. Because of that, fractionation and complementary strategies are required for its study. The MS analysis of complex samples may result in up to 100,000-peptide spectra that cannot be easily analyzed with standard procedures. Therefore, proteomics, more than other -omics, needs a dry lab, time, and an effort in data mining.As main conclusion, it can be stated that proteomics is in its beginnings. It is starting to make important contributions to a proper gene annotation, identification, and characterization of gene products or protein species and to the knowledge of living organisms, having also an enormous application potential to translational research. However, and despite its great potential, and as in any other experimental approach, it is far from being a Pandora's Box. In the case of plant research, the full potential of proteomics is quite far from being totally exploited, and second-, third-, and fourth-generation proteomics techniques are still of very limited use. Most of the plant proteomics papers so far published belong to the descriptive, subcellular, and comparative proteomics subgroup, mainly using a few experimental model systems-those whose genome has been sequenced-and being from a biological point of view quite descriptive and speculative. From now on we should put more emphasis on the study of posttranslational proteomics and interactomics, and move to targeted, hypothesis-driven approaches. Furthermore, and even more important, we should move to data validation through other -omics or classical biochemical strategies, in an attempt to get a deeper, real, and more accurate view and understanding of cell biology. In the modern Systems Biology concept, proteomics must be considered as a part of a global, multidisciplinary approach. Making biological sense of a proteomics experiment requires a proper experimental design, data validation, interpretation, and publication policy.