相关实验视频

Updated: May 28, 2025

Author Spotlight: Exploring the Mechanisms of MicroRNA Loading into Extracellular Vesicles in Cancer Progression

Author Spotlight: Exploring the Mechanisms of MicroRNA Loading into Extracellular Vesicles in Cancer Progression

Published on: October 6, 2023

囊泡和非囊泡细胞外tRNA及其碎片之间的歧视

Mauricio Castellano¹, Marco Li Calzi², Maria Rosa Garcia²

¹Functional Genomics Laboratory, Institut Pasteur de Montevideo, Montevideo, Uruguay; Immunoregulation and Inflammation Laboratory, Institut Pasteur Montevideo, Montevideo, Uruguay; Analytical Biochemistry Unit, Center for Nuclear Research, School of Science, Universidad de la Republica, Montevideo, Uruguay.

Methods in enzymology

|February 14, 2025

概括

研究人员开发了分离细胞外囊泡 (EVs) 和非EVs的协议,这对于生物标志物发现和基本生物学中准确的tRNA和tRNA衍生小RNA (tDR) 分析至关重要.

科学领域:

细胞外RNA生物学
发现生物标志物的发现.
分子生物学分子生物学

背景情况:

细胞外囊泡 (EV) 和非囊泡部分含有丰富的转移RNA (tRNA) 和tRNA衍生小RNA (tDR).
对细胞外tRNA和tDRs的准确分析需要区分EV-bound和非膀性RNAs.
非膀性tRNA可以被核糖酶降解成tDRs,导致测序数据中的潜在交叉污染.

研究的目的:

提供可靠的方法来将EV与非膀外细胞载体分离.
为了实现对细胞外tRNA和tDR群体的精确分析,用于生物和诊断应用.
为了防止因污染而导致细胞外RNA测序数据的误解.

主要方法:

密度梯度分离用于高分辨率分离细胞外组件.
使用商业列进行尺寸排除色谱,以实现更快,更常规的分离.
开发和验证EV隔离和随后RNA分析的协议.

主要成果:

为了有效地将EV与非膀外细胞RNA载体分离,建立了两个不同的协议.
密度梯度分离提供了高分辨率,但耗时.
尺寸排除色谱提供了一个更快,更简单的方法,适合常规使用.

结论:

精确的EV分离对于可靠的细胞外tRNA和tDR分析至关重要.
开发的协议为研究细胞外RNA生物学和寻找新生物标志物的研究人员提供了有价值的工具包.
这些方法将提高细胞外样本中小RNA测序的准确性.

关键词:

这就是ExRNA.外基因组是外基因组的组成部分.细胞外的tDRs是什么细胞外囊泡中的细胞外囊泡.非膀性 exRNA 的 RNA.

更多相关视频

Extraction of Extracellular Vesicles from Whole Tissue

Extraction of Extracellular Vesicles from Whole Tissue

Published on: February 7, 2019

Single Extracellular Vesicle Transmembrane Protein Characterization by Nano-Flow Cytometry

Single Extracellular Vesicle Transmembrane Protein Characterization by Nano-Flow Cytometry

Published on: July 26, 2022

相关概念视频

Vesicular Tubular Clusters

Vesicular Tubular Clusters

After budding out from the ER membrane, some COPII vesicles lose their coat and fuse with one another to form larger vesicles and interconnected tubules called vesicular tubular clusters or VTCs. These clusters constitute a compartment at the ER-Golgi interface known as ERGIC (Endoplasmic Reticulum Golgi Intermediate Compartment). The ERGIC is a mobile membrane-bound cargo transport system that sorts proteins secreted from ER and delivers them to the Golgi.
With the help of motor proteins such...

Transfer RNA Synthesis

Transfer RNA Synthesis

One of the unique features of tRNA is the presence of modified bases. In some tRNAs, modified bases account for nearly 20% of the total bases in the molecule. Altogether, these unusual bases protect the tRNA from enzymatic degradation by RNases.
Each of these chemical modifications is carried by a specific enzyme, post-transcription. All of these enzymes have unique base and site-specificity. Methylation, the most common chemical modification, is carried by at least nine different enzymes, with...

Leaky Scanning

Leaky Scanning

During most eukaryotic translation processes, the small 40S ribosome subunit scans an mRNA from its 5' end until it encounters the first start AUG codon. The large 60S ribosomal subunit then joins the smaller one to initiate protein synthesis. The location of the translation initiation is largely determined by the nucleotides near the start codon as there may be multiple translation initiation sites present on the mRNA. Marilyn Kozak discovered that the sequence RCCAUGG (where R...

Nuclear Export of mRNA

Nuclear Export of mRNA

Before mRNAs are exported to the cytoplasm, it is crucial to check each mRNA for structural and functional integrity. Eukaryotic cells use several different mechanisms, collectively known as mRNA surveillance, to look for irregularities in mRNAs. Irregular or aberrant mRNA are rapidly degraded by various enzymes. If a defective mRNA escapes the surveillance, it would be translated into a protein which would either be non-functional or not function properly. One of the primary irregularities in...

Improving Translational Accuracy

Improving Translational Accuracy

Base complementarity between the three base pairs of mRNA codon and the tRNA anticodon is not a failsafe mechanism. Inaccuracies can range from a single mismatch to no correct base pairing at all. The free energy difference between the correct and nearly correct base pairs can be as small as 3 kcal/ mol. With complementarity being the only proofreading step, the estimated error frequency would be one wrong amino acid in every 100 amino acids incorporated. However, error frequencies observed in...

Termination of Translation

Termination of Translation

The large ribosomal subunit has several important structures essential to translation. These include the peptidyl transferase center (PTC) - which is the site where the peptide bond is formed - and a large, internal, water-filled tube through which the nascent polypeptide moves. This latter structure is called the Peptide Exit Tunnel, and it begins at the PTC and spans the body of the large ribosomal subunit. During translation, as the nascent polypeptide chain is synthesized, it passes through...