کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1176580 | 961862 | 2007 | 14 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Monitoring repair of DNA damage in cell lines and human peripheral blood mononuclear cells
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کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
شیمی
شیمی آنالیزی یا شیمی تجزیه
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چکیده انگلیسی
We introduce a method to follow DNA repair that is suitable for both clinical and laboratory samples. An episomal construct with a unique 8-oxoguanine (8-oxoG) base at a defined position was prepared in vitro using single-stranded phage harboring a 678-bp tract from exons 5 to 9 of the human P53 gene. Mixing curve experiments showed that the real-time PCR method has a linear response to damage, suggesting that it is useful for DNA repair studies. The episomal construct with a unique 8-oxoG base was introduced into AD293 cells or human peripheral blood mononuclear cells, and plasmids were recovered as a function of time. The quantitative real-time PCR assay demonstrated that repair of the 8-oxoG was 80% complete in less than 48Â h in AD293 cells. Transfection of small interfering RNAs down-regulated OGG1 expression in AD293 cells and reduced the repair of 8-oxoG to 30%. Transfection of the episome into unstimulated white blood cells showed that 8-oxoG repair had a half-life of 2 to 5Â h. This method is a rapid, reproducible, and robust way to monitor repair of specific adducts in virtually any cell type.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Analytical Biochemistry - Volume 365, Issue 2, 15 June 2007, Pages 246-259
Journal: Analytical Biochemistry - Volume 365, Issue 2, 15 June 2007, Pages 246-259
نویسندگان
Hyun-Wook Lee, Hae-Jung Lee, Chong-mu Hong, David J. Baker, Ravi Bhatia, Timothy R. O'Connor,