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[利用模板色谱法从DNA部分水解产物中分离寡核苷酸]

[Isolation of oligonucleotides from partial hydrolysates of DNA using template chromatography].

作者信息

Schott H, Schrade H, Watzlawick H

出版信息

J Chromatogr. 1984 Mar 2;285(2):343-63. doi: 10.1016/s0021-9673(01)87775-x.

Abstract

Purine oligonucleotides are adsorbed at 0 degree C on poly(vinyl alcohol)-p(dC)n-DEAE-cellulose, and pyrimidine oligonucleotides on oligo(guanylic acid) gel according to the base-pairing mechanism, if their sequences contain at least three or more homologous, consecutive guanylic or cytidylic moieties. By increasing the temperature all base-paired oligonucleotides are desorbed. With this template chromatography mixtures of defined purine- or pyrimidine oligonucleotides could be isolated from fractionated partial hydrolyzates of herring sperm DNA. Afterwards these mixtures are rechromatographed on QAE-Sephadex and/or Nucleosil C18. Using this approach oligonucleotides up to nine monomer units can be isolated either as single substances or as mixtures with defined composition on a preparative scale, which would be not possible with the already existing separation procedures when partial hydrolyzates of herring sperm DNA as starting material are used. Purity and sequence of the isolated oligonucleotides are determined by the "fingerprint" method.

摘要

如果嘌呤寡核苷酸的序列包含至少三个或更多同源的、连续的鸟苷酸或胞苷酸部分,那么在0℃时,它们会根据碱基配对机制吸附在聚乙烯醇-p(dC)n-二乙氨基乙基纤维素上;嘧啶寡核苷酸则会吸附在寡聚(鸟苷酸)凝胶上。通过升高温度,所有碱基配对的寡核苷酸都会解吸。利用这种模板色谱法,可以从鲱鱼精DNA的分级部分水解产物中分离出特定的嘌呤或嘧啶寡核苷酸混合物。之后,将这些混合物在QAE-葡聚糖凝胶和/或硅胶C18上再次进行色谱分离。使用这种方法,可以在制备规模上以单一物质或具有确定组成的混合物形式分离出多达九个单体单元的寡核苷酸,而当使用鲱鱼精DNA的部分水解产物作为起始材料时,现有的分离程序无法做到这一点。分离出的寡核苷酸的纯度和序列通过“指纹”法确定。

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