Improvements in Pyrosequencing technology by employing Sequenase polymerase
2004 (English)In: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 330, no 2, 272-280 p.Article in journal (Refereed) Published
Pyrosequencing is a DNA sequencing technique based on the bioluminometric detection of inorganic pyrophosphate, which is released when nucleotides are incorporated into a target DNA. Since the technique is based on an enzymatic cascade, the choice of enzymes is a critical factor for efficient performance of the sequencing reaction. In this study we have analyzed the performance of an alternative DNA polymerase, Sequenase, on the sequencing performance of the Pyrosequencing technology. Compared to the Klenow fragment of DNA polymerase I, Sequenase could read through homopolymeric regions with more than five T bases. In addition, Sequenase reduces remarkably interference from primer-dimers and loop structures that give rise to false sequence signals. By using Sequenase, synchronized extensions and longer reads can be obtained on challenging templates, thereby opening new avenues for applications of Pyrosequencing technology.
Place, publisher, year, edition, pages
Academic Press, 2004. Vol. 330, no 2, 272-280 p.
pyrosequencing technology, sequenase, primer-dimers, loop structures, DNA sequencing
Biochemistry and Molecular Biology Chemical Sciences
IdentifiersURN: urn:nbn:se:umu:diva-85453DOI: 10.1016/j.ab.2004.03.018ISI: 000222424800012PubMedID: 15203333OAI: oai:DiVA.org:umu-85453DiVA: diva2:693578