Thymidine analogues for tracking DNA synthesis.
Thymidine analogues for tracking DNA synthesis.
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DOI：
10.3390/molecules16097980
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发表时间：
2011-09-15
期刊：
影响因子：
--
通讯作者：
Meedeniya AC
中科院分区：
文献类型：
--
作者：
Cavanagh BL;Walker T;Norazit A;Meedeniya AC
复制细胞在高度调控的细胞周期s期进行DNA合成。嘧啶脱氧核苷胸苷的类似物可以插入到复制的DNA中，有效地标记分裂的细胞，使其具有特征。氚化胸腺嘧啶在放射自显影技术上要求很高，并被5-溴-2-脱氧尿嘧啶（BrdU）和相关的卤化类似物所取代，可通过抗体检测。它们的检测需要DNA的变性，这往往限制了调查的结果。尽管存在这些局限性，但仅BrdU已在超过20,000项已审查的生物医学研究中用于靶向新合成的DNA。胸腺嘧啶类似物5-乙基-2 ' -脱氧尿嘧啶（EdU）是“标记DNA合成”的最新突破。利用荧光叠氮探针和铜催化的“Huisgen反应”（1,3-偶极环加成或“click化学”）很容易检测到EdU中的炔基。这种快速的两步生物标记方法允许细胞内DNA的标记和成像，同时保持细胞的结构和分子完整性。EdU的生物正交检测允许其应用于比以前更多的实验分析，而不是其他“非自然碱基”。这些包括在多个领域的生理、解剖和分子生物学实验，包括干细胞研究、癌症生物学和寄生虫学。EdU及其相关分子在生物医学研究中的全部潜力仍有待探索。
Replicating cells undergo DNA synthesis in the highly regulated, S-phase of the cell cycle. Analogues of the pyrimidine deoxynucleoside thymidine may be inserted into replicating DNA, effectively tagging dividing cells allowing their characterisation. Tritiated thymidine, targeted using autoradiography was technically demanding and superseded by 5-bromo-2-deoxyuridine (BrdU) and related halogenated analogues, detected using antibodies. Their detection required the denaturation of DNA, often constraining the outcome of investigations. Despite these limitations BrdU alone has been used to target newly synthesised DNA in over 20,000 reviewed biomedical studies. A recent breakthrough in “tagging DNA synthesis” is the thymidine analogue 5-ethynyl-2′-deoxyuridine (EdU). The alkyne group in EdU is readily detected using a fluorescent azide probe and copper catalysis using ‘Huisgen’s reaction’ (1,3-dipolar cycloaddition or ‘click chemistry’). This rapid, two-step biolabelling approach allows the tagging and imaging of DNA within cells whilst preserving the structural and molecular integrity of the cells. The bio-orthogonal detection of EdU allows its application in more experimental assays than previously possible with other “unnatural bases”. These include physiological, anatomical and molecular biological experimentation in multiple fields including, stem cell research, cancer biology, and parasitology. The full potential of EdU and related molecules in biomedical research remains to be explored.
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