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KMID : 1034820080040030246
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Molecular & Cellular Toxicology
2008 Volume.4 No. 3 p.246 ~ p.252
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Comparison of Non-amplified and Amplified DNA Preparation Methods for Array-comparative Gnomic Hybridization Analysis
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Joo Hong-Jin
Jung Seung-Hyun
Yim Seon-Hee
Kim Tae-Min
Xu Hai-Dong
Shin Seung-Hun
Kang Hyun-Mi
Chung Yeun-Jun
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Abstract
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Tumor tissue is usually contaminated by normal tissue components, which reduces the sensitivity of analysis for exploring genetic alterations. Although microdissection has been adopted to minimize the contamination of tumor DNA with normal cell components, there is a concern over the amount of microdissected DNA not enough to be applied to array-CGH reaction. To amplify the extracted DNA, several whole genome amplification (WGA) methods have been developed, but objective comparison of the array-CGH outputs using different types of WGA methods is still scarce. In this study, we compared the performance of non-amplified microdissected DNA and DNA amplified in 2 WGA methods such as degenerative oligonucleotide primed (DOP)-PCR, and multiple strand displacement amplification (MDA) using Phi 29 DNA polymerase. Genomic DNA was also used to make a comparison. We applied those 4 DNAs to whole genome BAC array to compare the false positive detection rate (FPDR) and sensitivity in detecting copy number alterations under the same hybridization condition. As a result microdissected DNA method showed the lowest FPDR and the highest sensitivity. Among WGA methods, DOP-PCR amplified DNA showed better sensitivity but similar FPDR to MDA-amplified method. These results demonstrate the advantage and applicability of microdissection for array-CGH analysis, and provide useful information for choosing amplification methods to study copy number alterations, especially based on precancerous and microscopically invaded lesions.
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KEYWORD
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Array-CGH (Comparative Genomic Hybridization), Microdissection, DOP-PCR (Degenerated Oligonucleotide-Primed PCR), Phi 29 DNA polymerase
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