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Complete protease inhibitor tablets, EDTA-free

cOmpleteTM protease inhibitor cocktail

Company: Roche Diagnostics
Catalog#: 11873580001
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Purification of Soluble Recombinant Human Tau Protein from Bacteria Using Double-tag Affinity Purification
Author:
Date:
2018-11-20
[Abstract]  Dysfunction of the microtubule-associated protein Tau (encoded by the MAPT gene) has been implicated in more than twenty neurodegenerative diseases, including Alzheimer’s. As such, the physiological and disease-relevant functions of Tau have garnered great interest in the research community. One barrier hampering investigations into the functions of Tau and the generation of pharmacological agents targeting Tau has been the difficulty of obtaining soluble Tau protein in purified form. Here, we describe a protocol that uses dual affinity tag purification to selectively purify soluble recombinant Tau protein from bacteria that is functionally active for downstream applications including immunization, microtubule binding assays, and protein-protein interaction studies. [摘要]  微管相关蛋白Tau(由 MAPT 基因编码)的功能障碍已经涉及20多种神经退行性疾病,包括阿尔茨海默病。 因此,Tau的生理和疾病相关功能引起了研究界的极大兴趣。 妨碍对Tau功能的研究和产生靶向Tau的药理学试剂的一个障碍是难以获得纯化形式的可溶性Tau蛋白。 在这里,我们描述了一种方案,该方案使用双亲和标签纯化从细菌中选择性纯化可溶性重组Tau蛋白,所述细菌对于下游应用具有功能活性,包括免疫,微管结合测定和蛋白质 - 蛋白质相互作用研究。
【背景】Tau传统上被定义为微管结合蛋白;然而,在人类疾病中,Tau可以与轴突微管分离并错误定位到其他神经元区室,包括体细胞,树突和突触,其中与非微管蛋白和结构的相互作用驱动神经元功能障碍(Iqbal et al。 ,2016; Wang和Mandelkow,2016; Zhou et al。,2017; McInnes et al。,2018)。尽管神经原纤维缠结形式的Tau聚集体通常存在于死后患病的脑组织中,但研究表明,可溶性Tau,而不是聚集的Tau,是神经元功能障碍的主要原因(Crimins et al。,2012 ; Polydoro et al。,2014; Koss et al。,2016)。因此,研究Tau在疾病中的可溶性功能,例如鉴定蛋白质 - ...

RNA Immunoprecipitation (RIP) Sequencing of Pri-miRNAs Associated with the Dicing Complex in Arabidopsis
Author:
Date:
2018-07-05
[Abstract]  RNA immunoprecipitation (RIP) is an antibody-based technique used to map in vivo RNA-protein interactions. DBR1, an RNA debranching enzyme, is responsible for the debranching of lariat RNA, for the degradation and turnover of lariat RNAs. It is well known that primary miRNA (Pri-miRNA) is recognized and further processed into mature miRNA by the Dicing complex mainly composed of DCL1 and HYL1. Due to the low abundance of pri-miRNAs, RIP followed qRT-PCR has been widely used to evaluate the binding efficiency of the Dicing complex with pri-miRNAs in previous studies. Therefore, the genome-wide evaluation of the Dicing complex with pri-miRNAs is lacking. With the improvement of high-throughput sequencing technologies, we successfully used RIP-seq to compare the binding efficiency ... [摘要]  RNA免疫沉淀(RIP)是一种基于抗体的技术,用于绘制体内 RNA-蛋白质相互作用。 DBR1是一种RNA脱支酶,负责套索RNA的脱支,用于套索RNA的降解和转换。众所周知,主要miRNA(Pri-miRNA)被主要由DCL1和HYL1组成的切割复合物识别并进一步加工成成熟miRNA。由于pri-miRNA的丰度较低,RIP随后的qRT-PCR已被广泛用于评估切割复合物与pri-miRNA在先前研究中的结合效率。因此,缺乏对具有pri-miRNA的切割复合物的全基因组评估。随着高通量测序技术的改进,我们成功地使用RIP-seq比较了Dicing复合物与野生型和 dbr1-2 突变体之间的pri-miRNA的结合效率。 。在该方案中,我们提供了在两种不同基因型之间的HYL1-YFP和DCL1-YFP转基因植物中使用GFP捕获珠的RIP-seq的详细描述。该方法可用于评估pri-miRNA与拟南芥中的切割复合物的结合,并且它可以应用于植物中的其他RNA结合蛋白。

Analysis of Autophagic Activity Using ATG8 Lipidation Assay in Arabidopsis thaliana
Author:
Date:
2018-06-20
[Abstract]  As a fundamental metabolic pathway to degrade and recycle cellular cargos, autophagy is highly induced upon stress, starvation and senescence conditions in plants. A double-membrane structure named autophagosome will form during this process for cargo sequestration and delivery into the vacuole.

A number of regulators have been characterized in plants, including the autophagy-related (ATG) proteins and other plant-specific proteins. Among them, ATG8 will undergo a lipidation process to become a membrane-bound ATG8-phosphatidylethanolamine form and mark the growing autophagosomal membrane as well as the completed autophagosome. Therefore, ATG8 has been regarded as a marker for autophagosomes; and biochemical detection of the membrane-associated form of ATG8 is used as one of ...
[摘要]  作为降解和回收细胞货物的基本代谢途径,自噬在植物的胁迫,饥饿和衰老条件下被高度诱导。 在这个过程中,将形成一个称为自噬体的双膜结构,用于货物隔离和输送到液泡中。

已经在植物中表征了许多调控因子,包括自噬相关(ATG)蛋白和其他植物特异性蛋白。 其中,ATG8将经历脂化过程以成为膜结合的ATG8-磷脂酰乙醇胺形式并标记日益增长的自噬体膜以及完成的自噬体。 因此,ATG8被认为是自噬体的标志; 并且膜结合形式的ATG8的生物化学检测被用作测量自噬活性的主要方法之一。 在这里,我们描述了使用拟南芥幼苗检测ATG8-PE形式的ATG8脂化测定法。

【背景】自噬是调节受损细胞器的大量降解和不需要的细胞内容物的基本代谢过程。在自噬过程中,称为自噬体的双膜结构将形成并将货物递送到液泡中以降解。自噬相关蛋白(ATG)需要调节自噬活性(Liu and ...

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