Microarrays Research Today is a free monthly online journal that collates and summarizes the latest research about Microarrays, including details on experiments, designs, statistics, analysis, software.

Microarray-based DNA resequencing using 3' blocked primers.

Sram J, Sommer SS, Liu Q

Department of Molecular Genetics, City of Hope National Medical Center, Duarte, CA 91010, USA.

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.

Published 1 February 2008 in Anal Biochem, 374(1): 41-7.
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