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P20/200/1000 Barrier pipette tips

Company: Genesee Scientific
Catalog#: 24-412
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Using RNA Sequencing and Spike-in RNAs to Measure Intracellular Abundance of lncRNAs and mRNAs
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Date:
2020-10-05
[Abstract]  Long noncoding RNAs (lncRNAs) play essential roles in normal physiology and in disease but their mechanisms of action can be challenging to identify. For mechanistic studies, it is often useful to know a lncRNA’s intracellular abundance, i.e., approximately how many molecules of the lncRNA are present in a typical cell of a cell-type of interest. At least two approaches have been used to approximate lncRNA intracellular abundance: single-molecule sensitivity RNA fluorescence in situ hybridization (smFISH) and single-gene, calibrated reverse-transcription followed by quantitative PCR (RT-qPCR). However, like all experimental approaches, these methods have their limitations. smFISH, when analyzed using diffraction-limited microscopy, can underestimate intracellular ... [摘要]  [摘要]长非编码RNA(lncRNA)在正常生理和疾病中起着至关重要的作用,但其作用机理可能难以鉴定。对于机理研究,了解lncRNA的胞内丰度(即在目标细胞类型的典型细胞中大约存在多少个lncRNA分子)通常很有用。至少两种方法已用于估算lncRNA细胞内丰度:单分子敏感性RNA荧光原位杂交(smFISH ...

Mutant Huntingtin Secretion in Neuro2A Cells and Rat Primary Cortical Neurons
Author:
Date:
2018-01-05
[Abstract]  Quantitative analysis of proteins secreted from the cells poses a challenge due to their low abundance and the interfering presence of a large amount of bovine serum albumin (BSA) in the cell culture media. We established assays for detection of mutant huntingtin (mHtt) secreted from Neuro2A cell line stably expressing mHtt and rat primary cortical neurons by Western blotting. Our protocol is based on reducing the amounts of BSA in the media while maintaining cell viability and secretory potential, and concentrating the media prior to analysis by means of ultrafiltration. [摘要]  由细胞分泌的蛋白质的定量分析由于它们的丰度低和在细胞培养基中干扰大量牛血清白蛋白(BSA)的存在而提出挑战。 我们建立检测突变亨廷顿蛋白(mHtt)检测分泌Neuro2A细胞系稳定表达mHtt和大鼠原代皮层神经元蛋白质印迹。 我们的方案是基于降低培养基中BSA的量,同时保持细胞活力和分泌潜能,并在通过超滤分析之前浓缩培养基。


【背景】许多蛋白质通过各种分泌途径从细胞分泌到细胞外环境中。这些途径包括在ER-高尔基体 - 质膜途径之后的常规分泌途径(Lee等,2004)和多个非常规途径,例如溶酶体胞吐作用,穿过质膜的易位和外泌体以及胞外体释放(Zhang和Schekman,2013)。为了研究这些途径,经常需要分析培养细胞分泌的蛋白质进入培养基。蛋白质可以游离形式分泌,也可以与胞膜结构如胞外体和外泌体结合(Zhang and ...

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