Method for determining the efficiency of double-stranded DNA double labeling with a fluorescent dye, fluorescence correlation spectroscopy and its application in the analysis of biochemical

摘要

the subject invention is a designation output double labelling dwuniciowego dna fluorescent dye using fluorescence correlation spectroscopy, comprising the following steps (a), where the first sample prepared by hot soak, and then was the lne cooling mixtures of two complementary dna jednoniciowych: of ssdna and of ssdna bthe fluorescent dye, and (b) preparing a second sample by hot soak, and then cooling the mixture composed of nieznakowanego jednoniciowego ssdna and nieznakowanego jednoniciowego ssdna b, and of jednoniciowego ssdna and of jednoniciowego ssdna b ba rwnikiem fluorescent.while in both samples concentration and quantity of all the ingredients used are the same; and (c) shall be measured separately for both these samples using fluorescence correlation spectroscopy as a result of this measurement is obtained autokorelacyjnych curves functions g (?) where? mean residence time of particles in the konfokalnym,with which it curves read parameters values g1, g2 (0) (0) for each of the functions autokorelacyjnych samples at point r = 0, and the value of trypletowych p and p2, and (d) is calculated (0) values ga and gb (0), and then (e) determines the efficiency x dou u00f3jnego marking dwuniciowego dnaon the basis of obtained from the measurement in the stage (c) and (d) values calculated in the stage, with the value of x, which is a fraction of the determining efficiency double labelling dwuniciowego dna.the invention also includes the use of dual labelling dna designated, the above described way in quantitative analysis of biochemical, especially in the analysis of enzyme activity, and to verify the measured value efficiency double labelling dna as well as in bad aniach cuts dna chain.