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一种用于选择性检测酵母线粒体DNA的荧光细胞化学方法

[A fluorescent-cytochemical method for selective detection of yeast mitochondrial DNA].

作者信息

Poglazova M N, Mashkovtseva A V

出版信息

Mikrobiologiia. 1990 Nov-Dec;59(6):1024-31.

PMID:2087204
Abstract

Yeast mitochondrial DNA (mDNA) can selectively be detected using a specific dye (DAPI). Nuclear DNA (nDNA) was stained along with mDNA only in three out of the fifteen studied yeast strains. Visualisation with a luminescent microscope showed that mDNA content varied among different yeast species as well as the size and shape of fluorescent mitochondrial nuclei. Intensive nDNA fluorescence was detected when a fixed specimen was treated with DAPI. Under these conditions, mDNA was revealed only in yeast cells with its high content. The process of fixation was shown to interfere with the integrity of mitochondria. It is also possible that the structure of DNA was modified to affect its interaction with the dye and thus the level of fluorescence. The developed technique of selective mDNA staining and the experimental results make it possible to gain a more accurate quantitative information about DNA content at the cellular level using cyto- and spectrofluorimetric techniques. This study involves important aspects pertinent to the dye interaction with yeast nuclear and mitochondrial DNA, both native and subjected to different fixation procedures.

摘要

酵母线粒体DNA(mDNA)可使用特定染料(4',6-二脒基-2-苯基吲哚,DAPI)进行选择性检测。在所研究的15种酵母菌株中,只有3种菌株的核DNA(nDNA)与mDNA一起被染色。用荧光显微镜观察显示,mDNA含量在不同酵母物种之间存在差异,荧光线粒体核的大小和形状也有所不同。当用DAPI处理固定标本时,检测到强烈的nDNA荧光。在这些条件下,mDNA仅在mDNA含量高的酵母细胞中显现出来。结果表明,固定过程会干扰线粒体的完整性。也有可能是DNA结构发生了改变,从而影响其与染料的相互作用,进而影响荧光水平。所开发的选择性mDNA染色技术和实验结果使得利用细胞荧光和光谱荧光技术在细胞水平上获得更准确的DNA含量定量信息成为可能。本研究涉及与染料与酵母核DNA和线粒体DNA(包括天然的以及经过不同固定程序处理的)相互作用相关的重要方面。

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