Immobilization of DNA with nitrogen mustard-biotin conjugate for global epigenetic analysis
Immobilization of DNA with nitrogen mustard-biotin conjugate for global epigenetic analysis
复制标题
DOI：
10.1016/j.aca.2018.09.008
复制
发表时间：
2018-12-28
影响因子：
6.2
通讯作者：
Kurita, Ryoji
中科院分区：
文献类型：
--
作者：
Kojima, Naoshi;Suda, Tomomi;Kurita, Ryoji
我们报道了用酶联免疫吸附试验（ELISA）对基因组DNA中具有代表性的表观遗传修饰5-甲基胞嘧啶进行定量分析。我们合成了一种由氮芥和生物素组成的新型杂双功能连接分子，用于捕获生物传感器件表面的DNA。该分子可以成功地将基因组DNA固定在链霉亲和素包被的96孔微孔板上，然后用于免疫化学表观遗传评估。我们实现了基因组DNA样品中5-mC的灵敏定量检测。从我们的系统中获得的小鼠大脑和小鼠小肠基因组的CpG甲基化率分别为79%和82%。这些数字与先前报道的75-85%的甲基化率很好地一致，这是通过全基因组亚硫酸酯测序确定的。因此，目前使用我们的新型双功能连接分子的技术提供了一种快速，简单，廉价的表观遗传评估方法，而不需要任何传统的亚硫酸盐处理，聚合酶链反应（PCR）或测序。(C) 2018 Elsevier B.V.版权所有
We report the quantitative analysis of 5-methylcytosine, a representative epigenetic modification in genomic DNA, with an enzyme-linked immunosorbent assay (ELISA). We synthesized a novel hetero-bifunctional linker molecule consisting of nitrogen mustard and biotin to capture DNA on the surface of biosensing devices. The molecule can successfully immobilize genomic DNA on a streptavidin coated 96-well microplate, which was then employed for immunochemical epigenetic assessment. We achieved the sensitive and quantitative detection of 5-mC in genomic DNA samples. The CpG methylation ratios obtained from our system for mouse brain and mouse small intestine genomes were 79% and 82%, respectively. These numbers are in good agreement with the previously reported methylation ratio of 75-85%, which was identified by whole genome bisulfite sequencing. Accordingly, the present technology using our novel bifunctional linker molecule provides a fast, easy, and inexpensive method for epigenetic assessment, without the need for any conventional bisulfite treatment, polymerase chain reaction (PCR), or sequencing. (C) 2018 Elsevier B.V. All rights reserved.