کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
7681629 1495821 2013 5 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
A label-free signal amplification assay for DNA detection based on exonuclease III and nucleic acid dye SYBR Green I
موضوعات مرتبط
مهندسی و علوم پایه شیمی شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
A label-free signal amplification assay for DNA detection based on exonuclease III and nucleic acid dye SYBR Green I
چکیده انگلیسی
We have developed a new fluorescence method for specific single-stranded DNA sequences with exonuclease III (Exo III) and nucleic acid dye SYBR Green I. It is demonstrated by a reverse transcription oligonucleotide sequence (target DNA, 27 bases) of RNA fragment of human immunodeficiency virus (HIV) as a model system. In the absence of the target DNA, the hairpin-probe is in the stem-closed structure, the fluorescence of SYBR Green I is very strong. In the presence of the target DNA, the hairpin-probe hybridizes with the target DNA to form double-stranded structure with a blunt 3′-terminus. Thus, in the presence of Exo III, only the 3′-terminus of probe is subjected to digestion. Exo III catalyzes the stepwise removal of mononucleotides from this terminus, releasing the target DNA. The released target DNA then hybridizes with another probe, whence the cycle starts anew. The signal of SYBR Green I decreases greatly. This system provides a detection limit of 160 pM, which is comparable to the existing signal amplification methods that utilized Exo III as a signal amplification nuclease. Due to the unique property of Exo III, this method shows excellent detection selectivity for single-base discrimination. More importantly, superiors to other methods based on Exo III, these probes have the advantages of easier to design, synthesize, purify and thus are much cheaper and more applicable. This new approach could be widely applied to sensitive and selective nucleic acids detection.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Talanta - Volume 114, 30 September 2013, Pages 49-53
نویسندگان
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