| Synaptoneurosome Preparation from C57BL/6 Striata
|
|
Author:
Date:
2016-02-20
[Abstract] Activity-dependent local mRNA translation endows synapses to remodel their structure and function (Bramham and Wells, 2007). This process is tightly controlled by the state of phosphorylation of several components of the translational machinery including initiation factors and ribosomal proteins (Buffington et al., 2014). The present protocol describes a method to prepare striatal synaptoneurosomes, from adult mice, containing both pre- and postsynaptic elements in which the level of synaptic phospho-proteins can be quantified (Biever et al., 2015).
[摘要] 活动依赖的局部mRNA翻译赋予突触重塑其结构和功能（Bramham和Wells，2007）。 该过程由包括起始因子和核糖体蛋白在内的翻译机理的几个组分的磷酸化状态来严格控制（Buffington等，2014）。 本方案描述了一种从成年小鼠制备纹状体synaptoneurosomes的方法，其含有可以量化突触磷酸蛋白水平的突触前和突触后元件（Biever等，2015）。
|
|
|
| Virus-induced Gene Silencing (VIGS) in Barley Seedling Leaves
|
|
Author:
Date:
2015-06-20
[Abstract] Virus induced gene silencing (VIGS) is one of the most potent reverse genetics technologies for gene functional characterisation. This method exploits a dsRNA-mediated antiviral defence mechanism in plants. Using this method allows researchers to generate rapid phenotypic data in a relatively rapid time frame as compared to the generation of stable transformants. Here we describe a simple method for silencing a target gene in barley seedling leaves using vectors based on the Barley Stripe Mosaic Virus (BSMV).
[摘要] 病毒诱导的基因沉默（VIGS）是基因功能表征的最有效的反向遗传学技术之一。 这种方法利用dsRNA介导的抗病毒防御机制在植物中。 与稳定转化体的产生相比，使用该方法允许研究人员在相对快的时间框架中产生快速表型数据。 在这里我们介绍了一种简单的方法，使用基于大麦条纹花叶病毒（BSMV）的载体，在大麦幼苗叶片中沉默目标基因。
|
|
|
| Analysis of Protein Stability by the Cycloheximide Chase Assay
|
|
Author:
Shih-Han Kao, Wen-Lung Wang, Chi-Yuan Chen, Yih-Leong Chang, Yi-Ying Wu, Yi-Ting Wang, Shu-Ping Wang, Alexey I Nesvizhskii, Yu-Ju Chen, Tse-Ming Hong and Pan-Chyr Yang,
Date:
2015-01-05
[Abstract] Comparison of protein stability in eukaryotic cells has been achieved by cycloheximide, which is an inhibitor of protein biosynthesis due to its prevention in translational elongation. It is broadly used in cell biology in terms of determining the half-life of a given protein and has gained much popularity in cancer research. Here we present a full cycloheximide chase assay in our laboratory using a lung adenocarcinoma cell line, CL1-5, as a model.
[摘要] 真核细胞中蛋白质稳定性的比较已经通过放线菌酮实现，放线菌酮是蛋白质生物合成的抑制剂，由于其在翻译延伸中的预防。 它在确定给定蛋白质的半衰期方面广泛用于细胞生物学，并且已在癌症研究中获得广泛应用。 在这里，我们在我们的实验室使用肺腺癌细胞系CL1-5作为模型呈现完整的放线菌酮追踪测定法。
|
|
|