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结合寡核苷酸阵列的制备与高质量引物的合成。

Combining the preparation of oligonucleotide arrays and synthesis of high-quality primers.

作者信息

Weiler J, Hoheisel J D

机构信息

Molecular-Genetic Genome Analysis, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 506, Heidelberg, D-69120, Germany.

出版信息

Anal Biochem. 1996 Dec 15;243(2):218-27. doi: 10.1006/abio.1996.0509.

Abstract

Based on the oligomer-chip technology, oligonucleotide arrays were synthesized directly on polypropylene sheets by a modified phosphoramidite chemistry using beta-eliminating nucleobase-protecting groups in combination with a succinate solid-phase linker. This method decouples the oligonucleotide deprotection from the support cleavage procedure, in contrast to standard phosphoramidite chemistry. In addition to being reliable substrates for hybridization experiments, the arrays also serve as source for the isolation of individual oligonucleotides. Technically, this allowed for a direct control of the quality of the arrayed oligomers. The released compounds were sufficient in amount and purity to work without further purification in PCR and DNA-sequencing reactions, with the results being identical to controls with commercially obtained primer molecules. Consequences for oligomer-chip hybridization procedures, the applicability of such hybrid-function arrays in, for example, diagnostics or comparative biology, and developments toward parallel primer synthesis are discussed.

摘要

基于寡聚物芯片技术,通过使用β-消除碱基保护基团与琥珀酸酯固相连接体相结合的改良亚磷酰胺化学方法,在聚丙烯片材上直接合成寡核苷酸阵列。与标准亚磷酰胺化学方法相比,该方法将寡核苷酸脱保护与载体切割过程分离。这些阵列除了是可靠的杂交实验底物外,还可作为分离单个寡核苷酸的来源。从技术上讲,这使得能够直接控制阵列寡聚物的质量。所释放的化合物在数量和纯度上足以在PCR和DNA测序反应中无需进一步纯化即可使用,结果与使用商业获得的引物分子的对照相同。文中还讨论了寡聚物芯片杂交程序的影响、此类杂交功能阵列在例如诊断或比较生物学中的适用性以及平行引物合成的进展。

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