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Investigation of genomic DNA methylation by ultraviolet resonant Raman spectroscopy.
Journal of Biophotonics ( IF 2.0 ) Pub Date : 2020-07-29 , DOI: 10.1002/jbio.202000150 Francesco D'Amico 1 , Paolo Zucchiatti 1, 2, 3 , Katia Latella 1, 4 , Maria Pachetti 1, 2 , Alessandro Gessini 1 , Claudio Masciovecchio 1 , Lisa Vaccari 1 , Lorella Pascolo 5
Journal of Biophotonics ( IF 2.0 ) Pub Date : 2020-07-29 , DOI: 10.1002/jbio.202000150 Francesco D'Amico 1 , Paolo Zucchiatti 1, 2, 3 , Katia Latella 1, 4 , Maria Pachetti 1, 2 , Alessandro Gessini 1 , Claudio Masciovecchio 1 , Lisa Vaccari 1 , Lorella Pascolo 5
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Cytosine plays a preeminent role in DNA methylation, an epigenetic mechanism that regulates gene expression, the misregulation of which can lead to severe diseases. Several methods are nowadays employed for assessing the global DNA methylation levels, but none of them combines simplicity, high sensitivity, and low operating costs to be translated into clinical applications. Ultraviolet (UV) resonant Raman measurements at excitation wavelengths of 272 nm, 260 nm, 250 nm, and 228 nm have been carried out on isolated deoxynucleoside triphosphates (dNTPs), on a dNTP mixture as well as on genomic DNA (gDNA) samples, commercial from salmon sperm and non‐commercial from B16 murine melanoma cell line. The 228 nm excitation wavelength was identified as the most suitable energy for enhancing cytosine signals over the other DNA bases. The UV Raman measurements performed at this excitation wavelength on hyper‐methylated and hypo‐methylated DNA from Jurkat leukemic T‐cell line have revealed significant spectral differences with respect to gDNA isolated from salmon sperm and mouse melanoma B16 cells. This demonstrates how the proper choice of the excitation wavelength, combined with optimized extraction protocols, makes UV Raman spectroscopy a suitable technique for highlighting the chemical modifications undergone by cytosine nucleotides in gDNA upon hyper‐ and hypo‐methylation events.
中文翻译：
通过紫外线共振拉曼光谱研究基因组DNA甲基化。
胞嘧啶在DNA甲基化中起着重要作用，DNA甲基化是一种调节基因表达的表观遗传机制，其表达异常会导致严重的疾病。如今，已经采用了几种方法来评估总体DNA甲基化水平，但是没有一种方法能够将简单，高灵敏度和低运行成本结合在一起，转化为临床应用。已在分离的脱氧核苷三磷酸（dNTP），dNTP混合物以及基因组DNA（gDNA）样品上进行了272 nm，260 nm，250 nm和228 nm激发波长的紫外（UV）共振拉曼测量，鲑鱼精液可商业获得，B16鼠黑色素瘤细胞系可商业获得。228 nm的激发波长被认为是增强其他DNA碱基胞嘧啶信号的最合适能量。在此激发波长下，对来自Jurkat白血病T细胞系的高甲基化和低甲基化DNA进行的紫外线拉曼测量显示，与分离自鲑鱼精子和小鼠黑素瘤B16细胞的gDNA相比，光谱存在显着差异。这证明了如何正确选择激发波长，并结合优化的提取方案，使紫外线拉曼光谱法成为一种合适的技术，可用来突出甲基化和甲基化过低事件中gDNA中胞嘧啶核苷酸所经历的化学修饰。
更新日期：2020-07-29
中文翻译：
通过紫外线共振拉曼光谱研究基因组DNA甲基化。
胞嘧啶在DNA甲基化中起着重要作用，DNA甲基化是一种调节基因表达的表观遗传机制，其表达异常会导致严重的疾病。如今，已经采用了几种方法来评估总体DNA甲基化水平，但是没有一种方法能够将简单，高灵敏度和低运行成本结合在一起，转化为临床应用。已在分离的脱氧核苷三磷酸（dNTP），dNTP混合物以及基因组DNA（gDNA）样品上进行了272 nm，260 nm，250 nm和228 nm激发波长的紫外（UV）共振拉曼测量，鲑鱼精液可商业获得，B16鼠黑色素瘤细胞系可商业获得。228 nm的激发波长被认为是增强其他DNA碱基胞嘧啶信号的最合适能量。在此激发波长下，对来自Jurkat白血病T细胞系的高甲基化和低甲基化DNA进行的紫外线拉曼测量显示，与分离自鲑鱼精子和小鼠黑素瘤B16细胞的gDNA相比，光谱存在显着差异。这证明了如何正确选择激发波长，并结合优化的提取方案，使紫外线拉曼光谱法成为一种合适的技术，可用来突出甲基化和甲基化过低事件中gDNA中胞嘧啶核苷酸所经历的化学修饰。
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