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The Efficacy of Hybridization Analysis of DNA Labeled with Red and Near-Infrared Cyanine Dyes
Biophysics Pub Date : 2021-04-27 , DOI: 10.1134/s000635092101005x
A. Yu. Ikonnikova , V. E. Shershov , Yu. V. Moroz , V. A. Vasiliskov , S. A. Lapa , R. A. Miftakhov , V. E. Kuznetsova , A. V. Chudinov , T. V. Nasedkina

We present a study of the sensitivity of a hybridization microarray analysis of DNA labeled with 2'‑deoxyuridine-5'-triphosphate derivatives that contain the Cy5 and Cy7 cyanine fluorophore dyes. Two Cy5-dUTP and two Cy7-dUTP derivatives that can be effectively incorporated into a newly synthesized DNA strand during PCR were chosen for this study. PCR products fluorescently labeled with different dyes were hybridized with a matrix of oligonucleotide probes immobilized in the microchip hydrogel cells. Fluorescent signals were recorded on a digital fluorescence microscope in the red (for Cy5-dUTP) and near-infrared (for Cy7-dUTP) range. Using the serial dilution method, the minimum DNA concentration was evaluated, which is necessary for genotyping by the specific interaction between the fluorescently labeled PCR product and oligonucleotide probes on the microarray. It has been shown that the fluorescently labeled Cy7-dUTP, unlike Cy5-dUTP, provides greater sensitivity of the hybridization analysis.



中文翻译:

红色和近红外花青染料标记的DNA杂交分析的功效

我们目前对杂交微阵列分析的敏感性进行了研究,该DNA用2'-脱氧尿苷-5'-三磷酸酯衍生物标记的DNA包含Cy5和Cy7花青荧光团染料。本研究选择了两个可以在PCR期间有效整合到新合成的DNA链中的Cy5-dUTP和两个Cy7-dUTP衍生物。将用不同染料荧光标记的PCR产物与固定在微芯片水凝胶细胞中的寡核苷酸探针基质杂交。在数字荧光显微镜上以红色(对于Cy5-dUTP)和近红外(对于Cy7-dUTP)范围记录荧光信号。使用系列稀释法评估了最低DNA浓度,这是通过荧光标记的PCR产物与微阵列上的寡核苷酸探针之间的特异性相互作用进行基因分型所必需的。已经表明,与Cy5-dUTP不同,荧光标记的Cy7-dUTP提供了更高的杂交分析灵敏度。

更新日期:2021-04-28
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