کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
2091667 1081743 2008 11 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Simultaneous quantification of multiple nucleic acid targets in complex rRNA mixtures using high density microarrays and nonspecific hybridization as a source of information
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی بیوتکنولوژی یا زیست‌فناوری
پیش نمایش صفحه اول مقاله
Simultaneous quantification of multiple nucleic acid targets in complex rRNA mixtures using high density microarrays and nonspecific hybridization as a source of information
چکیده انگلیسی

To date, it has been problematic to accurately quantify multiple nucleic acid sequences, representing microbial targets, in multi-target mixtures using oligonucleotide microarrays, primarily due to nonspecific target binding (i.e., cross-hybridization). While some studies ignore the effects of nonspecific binding, other studies have developed approaches to minimize nonspecific binding, such as physical modeling to design highly specific probes, subtracting nonspecific signal using mismatch probes, and/or removing nonspecific duplexes by scanning through a range of wash stringencies. We have developed an alternative approach that, in contrast to previous approaches, uses nonspecific target binding as a source of information. Specifically, the new approach uses hybridization patterns (fingerprints) to quantify specific nucleic acid targets in complex target mixtures. We evaluated the approach by mixing together in vitro transcribed 28S rRNA targets at varying concentrations (up to 1.0 nM), and hybridizing the 24 mixtures to microarrays (n = 3160 probes, in duplicate). Three independent Latin-square-designed experiments revealed accurate quantification of the targets. The regression between actual concentration of targets and those determined by the approach were highly positively correlated with high R2 values (e.g., R2 = 0.90, n = 6 targets; R2 = 0.84, n = 8 targets; R2 = 0.82, n = 10 targets).

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Microbiological Methods - Volume 75, Issue 1, September 2008, Pages 92–102
نویسندگان
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