کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1177282 | 961973 | 2008 | 7 صفحه PDF | دانلود رایگان |
To exceed the throughput and accuracy of conventional sequencing technologies, we tested a method (pyrophosphorolysis-activated polymerization [PAP]) of nucleic acid amplification that uses 3′ blocked primers (P∗s). As proof-of-principle, we resequenced a 20-bp region of the factor IX gene with a microarray of P∗s. P∗s discriminate 3′ end mismatches with ultra-high specificity as well as mismatches along their lengths with high specificity. We correctly identified two wild-type samples as well as all mismatches, including three single-base substitutions, one microdeletion, one microinsertion, and one heterozygous mutation. Despite limitations in the primer purity, the signal/noise ratio between the matched and mismatched P∗s sometimes exceeded 1000. Thus, PAP resequencing shows great potential for accurate and high-throughput microarray-based resequencing.
Journal: Analytical Biochemistry - Volume 374, Issue 1, 1 March 2008, Pages 41–47