| Pipecolic Acid Quantification Using Gas Chromatography-coupled Mass Spectrometry
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Author:
Date:
2020-12-05
[Abstract] Pipecolic acid (Pip), a non-proteinacious product of lysine catabolism, is an important regulator of immunity in plants and humans alike. For instance, Pip accumulation is associated with the genetic disorder Zellweger syndrome, chronic liver diseases, and pyridoxine-dependent epilepsy in humans. In plants, Pip accumulates upon pathogen infection and is required for plant defense. The aminotransferase ALD1 catalyzes biosynthesis of Pip precursor piperideine-2-carboxylic acid, which is converted to Pip via ornithine cyclodeaminase. A variety of methods are used to quantify Pip, and some of these involve use of expensive amino acid analysis kits. Here, we describe a simplified procedure for quantitative analysis of Pip from plant tissues. Pipecolic acid was extracted from leaf tissues along ...
[摘要] [摘要]胡椒酸（Pip）是赖氨酸分解代谢的非蛋白质产物，是植物和人类免疫力的重要调节剂。例如，Pip积累与人类遗传病Zellweger综合征，慢性肝病和吡ido醇依赖性癫痫有关。在植物中，Pip会在病原体感染时积聚，是植物防御所必需的。氨基转移酶ALD1催化Pip前体哌啶-2-羧酸的生物合成，后者通过鸟氨酸环脱氨酶转化为Pip。多种方法可用于定量Pip，其中一些方法涉及使用昂贵的氨基酸分析试剂盒。在这里，我们描述了从植物组织中定量定量分析的简化程序。P ipecolic酸是从叶组织中提取与内标正缬氨酸一起衍生与丙基氯甲酸酯和分析通过气相色谱-质谱联用选择性离子模式。此过程简单，经济且有效，并且不涉及同位素内标或多步衍生化。
关键词：胡椒酸GC-MS拟南芥植物防御氯甲酸丙酯
[背景]匹，非蛋白氨基酸赖氨酸的氨基酸衍生物，积聚在病原体感染的植物的本地和远端的组织和患者的体液与过氧化物酶体疾病（Schutgens等人，1986;尹和An，2010;Návarová等人。，2012;王等人，2018） ...
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| Rapid Generation of Human Neuronal Cell Models Enabling Inducible Expression of Proteins-of-interest for Functional Studies
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Author:
Date:
2020-05-05
[Abstract] CRISPR-Cas9 technology has transformed the ability to edit genomic sequences and control gene expression with unprecedented ease and scale. However, precise genomic insertions of coding sequences using this technology remain time-consuming and inefficient because they require introducing adjacent single-strand cuts through Cas9 nickase action and invoking the host-encoded homology-directed repair program through the concomitant introduction of large repair templates. Here, we present a system for the rapid study of any protein-of-interest in two neuronal cell models following its inducible expression from the human AAVS1 safe harbor locus. With lox-flanked foundation cassettes in the AAVS1 site and a tailor-made plasmid for accepting coding sequences-of-interest in ...
[摘要] [摘要] CRISPR-Cas9技术以前所未有的简便性和规模改变了编辑基因组序列和控制基因表达的能力。但是，由于需要引入相邻的单链，因此使用该技术进行精确的基因组编码插入仍然很耗时且效率低下。通过减少Cas9切口酶的作用并通过同时引入大型修复模板来调用宿主编码的同源性指导的修复程序。在此，我们提出了一种系统，用于在其诱导后的两个神经元细胞模型中快速研究任何目的蛋白该系统可从人类AAVS1 安全港基因座表达，在AAVS1 位点具有lox侧翼的基础盒和定制的质粒以接受感兴趣的编码序列，该系统使研究人员能够为诱导型产生自己的神经元细胞模型任何编码表达序列不到一个月的时间。由于可用性Preinserted 增强型绿色的Fluo 可以与目标蛋白质融合的最新蛋白质（EGFP）编码序列，该系统可帮助功能研究通过活细胞显微镜以及利用非常有效的可用性进行的相互作用组分析来跟踪目标蛋白质EGFP捕获矩阵。
[背景] ...
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