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通过化学降解对寡核苷酸进行非放射性自动测序。

Non-radioactive automated sequencing of oligonucleotides by chemical degradation.

作者信息

Ansorge W, Rosenthal A, Sproat B, Schwager C, Stegemann J, Voss H

机构信息

European Molecular Biology Laboratory, Heidelberg, FRG.

出版信息

Nucleic Acids Res. 1988 Mar 25;16(5):2203-6. doi: 10.1093/nar/16.5.2203.

DOI:10.1093/nar/16.5.2203
PMID:3357772
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC338209/
Abstract

A non-radioactive sequencing of fluorescently labelled oligonucleotides by solid-phase chemical degradation is described. Although non-radioactive methods have been reported for the dideoxy chain termination technique, such a method has not yet been developed for the chemical degradation sequencing of DNA fragments. A 21-mer fluorescein labelled M13 sequencing primer was sequenced in an on-line automated system in about 30 minutes. The fluorescent dye and its bond to the oligonucleotide were stable during the chemical reactions used for the base specific degradations. As the sequence is determined on-line during electrophoresis, reloading and running 10 fragments simultaneously allows us to use one gel for sequencing of about 50 different oligonucleotides.

摘要

本文描述了通过固相化学降解对荧光标记寡核苷酸进行非放射性测序的方法。虽然已经报道了用于双脱氧链终止技术的非放射性方法，但尚未开发出用于DNA片段化学降解测序的此类方法。一个21聚体荧光素标记的M13测序引物在在线自动化系统中约30分钟内完成了测序。在用于碱基特异性降解的化学反应过程中，荧光染料及其与寡核苷酸的连接是稳定的。由于序列在电泳过程中在线确定，同时重新加载并运行10个片段使我们能够用一块凝胶对约50种不同的寡核苷酸进行测序。

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本文引用的文献

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Nucleic Acids Res. 1984 Jun 11;12(11):4539-57. doi: 10.1093/nar/12.11.4539.
2
Solid-phase methods for sequencing of nucleic acids I. Simultaneous sequencing of different oligodeoxyribonucleotides using a new, mechanically stable anion-exchange paper.核酸测序的固相方法I. 使用一种新型机械稳定阴离子交换纸同时对不同寡脱氧核糖核苷酸进行测序
Nucleic Acids Res. 1985 Feb 25;13(4):1173-84. doi: 10.1093/nar/13.4.1173.
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Fluorescence detection in automated DNA sequence analysis.自动DNA序列分析中的荧光检测。
Nature. 1986;321(6071):674-9. doi: 10.1038/321674a0.
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A simple and rapid preparation of M13 sequencing templates for manual and automated dideoxy sequencing.一种用于手动和自动双脱氧测序的M13测序模板的简单快速制备方法。
Nucleic Acids Res. 1987 Jul 24;15(14):5507-16. doi: 10.1093/nar/15.14.5507.
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Automated DNA sequencing: ultrasensitive detection of fluorescent bands during electrophoresis.自动化DNA测序：电泳过程中荧光条带的超灵敏检测。
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