Chemical methods of DNA and RNA fluorescent labeling
Chemical methods of DNA and RNA fluorescent labeling
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DOI：
10.1093/nar/24.22.4535
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发表时间：
1996-11-15
影响因子：
14.9
通讯作者：
Mirzabekov, A
中科院分区：
文献类型：
--
作者：
Proudnikov, D;Mirzabekov, A
已经描述了几种用于DNA和RNA的荧光标记的方法，它们基于通过DNA的部分脱嘌呤或通过高碘酸钠氧化RNA中的3 '-末端核糖核苷来引入醛基。具有连接的肼基的荧光标记与醛基有效地偶联，并且通过用氰基硼氢化钠还原来稳定腙键。用乙二胺可以在脱嘌呤位点定量地裂解DNA，通过用氰基硼氢化钠还原来稳定脱嘌呤DNA或氧化RNA中的醛基与乙二胺之间的醛亚胺键，并且在这些位点引入的伯胺基团用于连接荧光染料的异硫氰酸酯或琥珀酰亚胺衍生物，荧光DNA标记可以在溶液中或在反相柱上进行。这些方法提供了简单、廉价的多个DNA标记和每个RNA引入一个荧光染料分子的方法，以及定量DNA片段化和每个片段掺入一个标记的方法，这些荧光团附着的方法被证明是有效的用于标记的RNA，DNA和DNA片段与寡核苷酸微芯片的杂交。
Several procedures have been described for fluorescent labeling of DNA and RNA, They are based on the introduction of aldehyde groups by partial depurination of DNA or oxidation of the 3'-terminal ribonucleoside in RNA by sodium periodate, Fluorescent labels with an attached hydrazine group are efficiently coupled with the aldehyde groups and the hydrazone bonds are stabilized by reduction with sodium cyanoborohydride, Alternatively, DNA can be quantitatively split at the depurinated sites with ethylenediamine, The aldimine bond between the aldehyde group in depurinated DNA or oxidized RNA and ethylenediamine is stabilized by reduction with sodium cyanoborohydride and the primary amine group introduced at these sites is used for attachment of isothiocyanate or succinimide derivatives of fluorescent dyes, The fluorescent DNA labeling can be carried out either in solution or on a reverse phase column, These procedures provide simple, inexpensive methods of multiple DNA labeling and of introducing one fluorescent dye molecule per RNA, as well as quantitative DNA fragmentation and incorporation of one label per fragment, These methods of fluorophore attachment were shown to be efficient for use in the hybridization of labeled RNA, DNA and DNA fragments with oligonucleotide microchips.