Evaluation of procedures for amplification of small-size samples for hybridization on microarrays

Various approaches have been developed for the preparation of samples for gene expression monitoring. For Affymetrix chips, a standard protocol is widely used; however, this is inefficient for small samples such as laser capture microdissections. Several amplification procedures for such samples alr...

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Bibliographic Details
Published in:Genomics (San Diego, Calif.) Calif.), 2004-03, Vol.83 (3), p.508-517
Main Authors: Klur, Sandra, Toy, Karen, Williams, Mickey P, Certa, Ulrich
Format: Article
Language:English
Subjects:
Animals
Biological and medical sciences
Brain - metabolism
Evaluation Studies as Topic
Female
Fundamental and applied biological sciences. Psychology
Gene Expression Profiling
Genes. Genome
Genetics of eukaryotes. Biological and molecular evolution
Laser capture microdissection
Mice
Microdissection - methods
Models, Biological
Molecular and cellular biology
Molecular genetics
Nucleic Acid Amplification Techniques - methods
Oligonucleotide Array Sequence Analysis - instrumentation
Oligonucleotide Array Sequence Analysis - methods
Oligonucleotide microarrays
Reproducibility of Results
Research Design
RNA amplification techniques
RNA, Complementary - metabolism
Sensitivity and Specificity
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Summary:Various approaches have been developed for the preparation of samples for gene expression monitoring. For Affymetrix chips, a standard protocol is widely used; however, this is inefficient for small samples such as laser capture microdissections. Several amplification procedures for such samples already exist, and our goal was to test two of them: the first is based on random PCR amplification, and the second, linear amplification, involves performing the standard protocol twice. We analyzed a dilution of a commercially available mouse brain total RNA preparation and microdissections from mouse hippocampus and striatum. We evaluated the quality of microarray data by analyzing several chip parameters and performing multiple comparisons. At the biological level, brain microdissections prepared with either method gave similar expression results. At the technical level, analysis of the commercial sample showed that random PCR amplification is more reproducible, requires smaller RNA input, and generates cRNA of higher quality than linear amplification.
ISSN:0888-7543
1089-8646
DOI:10.1016/j.ygeno.2003.09.005