کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
7232113 | 1470961 | 2015 | 6 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Site-specific immunochemical methylation assessment from genome DNA utilizing a conformational difference between looped-out target and stacked-in nontarget methylcytosines
دانلود مقاله + سفارش ترجمه
دانلود مقاله ISI انگلیسی
رایگان برای ایرانیان
کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
شیمی
شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله

چکیده انگلیسی
We report the sequence-selective immunochemical discrimination of methylcytosine from genomic DNA that we achieved by utilizing selective antibody binding to a looped-out methylcytosine in a bulge region and without using bisulfite treatment, a methylation-sensitive restriction enzyme, or PCR. First, we investigated the affinity of an anti-methylcytosine antibody for methylcytosine in full match, bulge, mismatch and abasic sites using surface plasmon resonance measurement and a microtiter plate to explore the differences as regards antibody binding to the target methylcytosine. The highest affinity, which was comparable to that in a single strand of DNA, was observed in the bulge region. In particular, no affinity was observed in a full match site. This is because there is no interaction such as hydrogen bond or Ï-Ï stacking for the bulged methylcytosine, thus enabling only the target in the bulge to be looped out. Methylated and unmethylated genomic DNA were blended to form a model DNA with which to assess the methylation ratio at a specific site. Fragmented DNA was hybridized with a biotinylated probe DNA, which has a sequence capable of forming a single base bulge at the target. The probe design is simple because it consists solely of the elimination of guanine paired with the target cytosine from a full match sequence. As a result, we successfully obtained a linear relationship (r2=0.9962) between the immunoassay signal and the methylation ratio of a specific site within 4Â h.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biosensors and Bioelectronics - Volume 70, 15 August 2015, Pages 366-371
Journal: Biosensors and Bioelectronics - Volume 70, 15 August 2015, Pages 366-371
نویسندگان
Ryoji Kurita, Hiroyuki Yanagisawa, Kyoko Yoshioka, Osamu Niwa,