Analysis of Sources of Error in Quantitation of Purified DNA Fragments and Unpurified PCR Products by DNA Microchip Electrophoresis |
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| Authors: | M R Mohamadi M Kataoka L Mahmoudian M Jabasini Y Shinohara Y Baba |
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| Institution: | (1) Department of Molecular and Pharmaceutical Biotechnology, Graduate School of Pharmaceutical Sciences, The University of Tokushima, Tokushima 770–8505, Japan;(2) The 21st Century COE Program, The University of Tokushima, Tokushima, Japan;(3) CREST, Japan Science and Technology Corporation, Tokushima, Japan;(4) Division of Gene Expression, Institute for Genome Research, The University of Tokushima, Japan;(5) Single-Molecule Bioanalysis Laboratory, National Institute of Advanced Industrial Science and Technology, Takamatsu, Japan;(6) Department of Applied Chemistry, Graduate School of Engineering Nagoya University, Nagoya 464–8603, Japan |
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| Abstract: | We have investigated the accuracy and reproducibility of DNA quantitation on the DNA Lab-Chip and the relationship of these to the size and concentration of the DNA fragments. We found that quantitation of small DNA fragments, i.e. less than 200 bp, suffers from high relative error which can be improved by using an internal standard of similar size to the sample. The effects of trace chloride ion on quantitation error and sensitivity of the DNA Lab-Chip were also studied, and it was revealed that 0.2 mM chloride ion reduces quantitation sensitivity by 30% and increases the relative error. We also studied the effects of purification on quantitation errors in analysis of PCR products from cloning vector pUC118 and showed that use of an unpurified sample reduces chip sensitivity by 25%.Dedicated to Professor K. Jinno on the occasion of his 60th birthday.Revised: 9 December 2004 and 17 January 2005 |
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| Keywords: | Electrophoresis Microchip construction DNA quantitation PCR product |
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