Detection method for quantifying global DNA methylation byfluorescence correlation spectroscopy
| Project/Area Number |
23701087
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| Research Category |
Grant-in-Aid for Young Scientists (B)
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| Allocation Type | Multi-year Fund |
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| Research Field |
Tumor diagnosis
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| Research Institution | Tokyo Medical University |
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Principal Investigator |
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| Project Period (FY) |
2011 – 2012
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| Project Status |
Completed (Fiscal Year 2012)
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| Budget Amount *help |
¥3,380,000 (Direct Cost: ¥2,600,000、Indirect Cost: ¥780,000)
Fiscal Year 2012: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
Fiscal Year 2011: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
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| Keywords | ゲノム / DNA メチル化 / 脱メチル化薬 / 1分子蛍光分析法 / MBD2 / メチル化DNA / DNA脱メチル化薬 / 一分子蛍光分析法 / 骨髄異形成症候群 |
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| Research Abstract |
A method for quantifying global DNA methylation using fluorescence correlation spectroscopy (FCS) has been established. The single-molecule methylation assay (SMMA) is based on two methodologies. One methodology, FCS, estimates the translational diffusion coefficient of molecules in solution, whereas the other methodology uses the high affinity of methyl-CpG-binding domain protein 2 (MBD2) to bind specifically to methylated DNA. We studied the specific binding rates of fluorescence-labeled MBD2 and methylated DNA from biological samples using the automated FCS system. Using a standard curve with methylated control DNA, we developed the SMMA index to assess the global DNA methylation level of the biological samples. A marked decrease in the SMMA index was observed when human leukemia cell lines (U937 and K562) were cultured with DNA demethylating agents. Our findings clearly indicate the applicability of SMMA as a simple and rapid tool for quantifying global DNA methylation. SMMA may prove useful for genome-wide comparative methylation analyses of malignancies and as an indicator of the demethylation effects of epigenetic drugs.
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Report
(3 results)
Research Products
(19 results)
