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Arrayed primer extension on in situ synthesized 5′ → 3′ oligonucleotides in microchannels

Paper ID Volume ID Publish Year Pages File Format Full-Text
33926 44998 2008 9 PDF Available
Title
Arrayed primer extension on in situ synthesized 5′ → 3′ oligonucleotides in microchannels
Abstract

Arrays of oligonucleotides synthesized in the 5′ → 3′ direction have potential benefit in several areas of life sciences research because the free 3′-end can be modified by enzymatic reactions. A Geniom One instrument (febit biomed GmbH, Germany), with integrated chip fabrication, multiplex primer extension, fluorescence imaging, and data analysis, was evaluated for studies of genomic variations. Microchannels used for the array synthesis in Geniom One were not optimized before for the APEX method and, as preliminary experiments demonstrated in this study, the signals were strongly affected by the speed of the process inside reaction channels. Using the two-compartment model (TCM), target binding to feature were quantitatively analyzed, revealing profound mass-transport limitations in the observed kinetics and enabling us to draw a series of physicochemical conclusions of the optimal set-up for the APEX reaction. Some kinetically relevant parameters such as target concentration, reaction time, and temperature were comprehensively analyzed. Finally, we applied the arrays and methods in a proof-of-principle experiment where 36 individuals were typed with 900 oligonucleotide probes (sense and antisense primers for 450 markers), using the ABCR gene as a test system. A new DNA analysis method for studies of genomic variation was developed using this all-in-one platform.

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Arrayed primer extension on in situ synthesized 5′ → 3′ oligonucleotides in microchannels
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Publisher
Database: Elsevier - ScienceDirect
Journal: New Biotechnology - Volume 25, Issues 2–3, October–December 2008, Pages 133–141
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us