DNA发夹结构在体外重新打开温度的测定。

Xuefeng Pan
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引用次数: 1

摘要

本文提出了一种基于引物延伸的DNA发夹结构重开温度测量方法。两个DNA寡核苷酸已被利用,并指定为引物1和2。引物1的5′端和3′端与发夹侧翼序列完全互补,用于评价引物延伸条件;引物2的3′端与DNA发夹茎竞争,用于检测DNA发夹重新打开温度。通过热变性和热再变性,在DNA模板上形成了单个的DNA发夹结构,这种发夹结构可以防止引物2的3′端与模板DNA一起退火,从而导致引物无法延伸。通过在不同的温度下孵育,DNA发夹结构可以在特定的温度下重新打开,引物延伸可以进行。这导致在琼脂糖凝胶上检测到双链DNA的出现。这个温度在这里被定义为发夹重开温度。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Determination of the reopening temperature of a DNA hairpin structure in vitro.

A novel method, based upon primer extension, has been developed for measuring the reopening temperature of a single type of DNA hairpin structure. Two DNA oligonucleotides have been utilized and designated as primers 1 and 2. Primer 1, with its 5- and 3'-termini fully complementary to the hairpin flanking sequences, was used to evaluate primer extension conditions, and primer 2, with its 3'-end competing with the DNA hairpin stem, was used to detect the DNA hairpin reopening temperature. A single DNA hairpin structure was formed on the DNA template by thermal denaturation and renaturation, and this hairpin structure was predicted to prevent the annealing of the 3'-end of primer 2 with the template DNA, which leads to no primer extension. By incubating at different temperatures, the DNA hairpin structure can be reopened at a particular temperature where the primer extension can be carried out. This resulted in the appearance of double-stranded DNA that was detected on an agarose gel. This temperature is defined here as the hairpin reopening temperature.

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