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• EU (5-ethynyl uridine)

EU (5-ethynyl uridine)

5-ethynyl uridine (EU) is a commonly used molecular biology reagent for RNA synthesis studies.

EU is readily taken up by living cells and phosphorylated through the pyrimidine salvage pathway. Generated 5-ethynyl uridine-5’-triphosphate is incorporated by RNA polymerases I, II, and III into de novo RNA instead of uridine, but not into DNA.

EU-labeled nascent cellular RNA can be detected quickly and with high sensitivity via Click Chemistry following fluorescent visualization. Alkyne group attached at the 5-position of uridine in modified RNA reacts with dye or biotin azides via Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC).

Labeled RNA can be detected with different methods, e.g. fluorescent microscopy or flow cytometry, which allows estimating transcriptional levels in the cells.

5-ethynyl uridine has an advantage over its analog, 5-bromouridine, because azide-containing dyes are very small in size and exhibit better membrane permeability compared to antibodies used for the detection of 5-bromouridine. Thus, the Сlick Сhemistry approach allows whole-mount staining of large samples like organs or tissue fragments.

5-乙炔基尿苷 (EU) 是用于 RNA 合成研究的常用分子生物学试剂。

EU 很容易被活细胞吸收并通过嘧啶补救途径磷酸化。生成的 5-乙炔基尿苷-5′-三磷酸被 RNA 聚合酶 I、II 和 III 整合到从头 RNA 中，而不是整合到 DNA 中。

EU 标记的新生细胞 RNA 可以在荧光可视化后通过 Click Chemistry 快速且高灵敏度地检测到。连接在修饰 RNA 中尿苷 5 位的炔基通过 Cu(I) 催化的叠氮化物-炔烃环加成 (CuAAC) 与染料或生物素叠氮化物反应。

标记的 RNA 可以用不同的方法检测，例如荧光显微镜或流式细胞术，可以估计细胞中的转录水平。

5-乙炔基尿苷优于其类似物 5-溴尿苷，因为与用于检测 5-溴尿苷的抗体相比，含叠氮化物的染料尺寸非常小并且表现出更好的膜渗透性。因此，Сlick Сhemistry 方法允许对器官或组织碎片等大样本进行整体染色。