Methods in molecular biology
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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. ⋯ 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.
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Multiple sequence alignment (MSA) is a fundamental and ubiquitous technique in bioinformatics used to infer related residues among biological sequences. Thus alignment accuracy is crucial to a vast range of analyses, often in ways difficult to assess in those analyses. ⋯ We outline a set of desirable characteristics for effective benchmarking, and evaluate each strategy in light of them. We conclude that there is currently no universally applicable means of benchmarking MSA, and that developers and users of alignment tools should base their choice of benchmark depending on the context of application-with a keen awareness of the assumptions underlying each benchmarking strategy.
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Targeted intrathecal (IT) drug delivery systems (IDDS) are an option in algorithms for the treatment of patients with moderate to severe chronic refractory pain when more conservative options fail. This therapy is well established and supported by several publications. ⋯ Recent technological advances, new therapeutic applications, reported complications, and the costs as well as maintenance required for this therapy require the need to stay up-to-date about new recommendations that may improve outcomes. This chapter reviews all technological issues regarding IDDS implantation with follow-up, and pharmacological recommendations published during recent years that provide evidence-based decision making process in the management of chronic pain and spasticity in patients.
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Metabolites, the chemical entities that are transformed during metabolism, provide a functional readout of cellular biochemistry that offers the best prediction of the phenotype and the nature of a disease. Mass spectrometry now allows thousands of metabolites to be quantitated. ⋯ These sophisticated statistical techniques are computationally intensive. This chapter reviews techniques applicable to metabolomics approaches to disease.
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N-methyl-D-aspartate receptors (NMDARs) are ionotropic glutamate receptors that are essential for synaptic plasticity, learning and memory. Dysfunction of NMDARs has been implicated in many nervous system disorders; therefore, pharmacological modulation of NMDAR activity has great therapeutic potential. However, given the broad physiological importance of NMDARs, modulating their activity often has detrimental side effects precluding pharmaceutical use of many NMDAR modulators. ⋯ Additionally, the effects of drugs on agonist-activated responses typically depend on duration of agonist exposure. To evaluate drug effects on synaptic transmission, an approach should be used that allows for activation of receptor responses as brief as those observed during synaptic transmission, both in the absence and presence of drug. To address these issues, we designed a fast perfusion system capable of (1) delivering brief (~5 ms) and consistent applications of glutamate to recombinant NMDARs of known subunit composition, and (2) easily and quickly (~5 s) changing between glutamate applications in the absence and presence of drug.