کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10559454 | 969627 | 2011 | 6 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Nanoliter droplet array for microRNA detection based on enzymatic stem-loop probes ligation and SYBR Green real-time PCR
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کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
شیمی
شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
In this paper, a nanoliter droplet array based on enzymatic stem-loop probes ligation and SYBR Green real-time PCR for quantification of microRNA was developed. By employing T4 RNA ligase 2 instead of T4 DNA ligase, we designed simplified stem-loop probes to perform microRNA-templated DNA ligation and reduced the non-specific ligation of T4 DNA ligase. SYBR green I dye was employed instead of TaqMan probes in present miniaturized real-time PCR systems. Specifically, we optimized the dosage of SYBR Green I dye in nanoliter droplet and verified the performance of this system by detecting synthetic mir-122 with a 6 logs dynamic range (from 1.5Â ÃÂ 105 to 1.5Â ÃÂ 1010 copies). Linear relationship of the standard curve (R2Â =Â 0.9997) and high PCR amplification efficiency (96.83%) were obtained under the optimized conditions. We detected the expression of mir-122 across five mouse tissues and the result was consistent with that TaqMan microRNA assay. We think this miniaturized real-time PCR platform reduced the detection cost considerably, thus showing the great potential to quantitative biology.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Talanta - Volume 85, Issue 4, 30 September 2011, Pages 1760-1765
Journal: Talanta - Volume 85, Issue 4, 30 September 2011, Pages 1760-1765
نویسندگان
Zeqi Yu, Ying Zhu, Yunxia Zhang, Juan Li, Qun Fang, Jianzhong Xi, Bo Yao,