International Mammalian Genome Society

logo18th International Mouse Genome Conference

17-22 October 2004, Seattle, USA


Carter MG 1, Sharov AA 1, VanBuren V 1, Dudekula DB 1, Carmack CE 2, Ko MSH 1

1 National Institute on Aging, Baltimore, MD, United States, 2 Agilent Technologies, Palo Alto, CA, United States

In an effort to enable more comprehensive expression profiling studies of mouse embryos and stem cells, we have created a 60-mer oligo microarray platform with coverage of all genes and transcripts identified in the NIA Mouse Gene Index 2.0.  The NIA Gene Index combines genome assembly sequence, public EST libraries, and human-curated genetic databases to form structure models of over 40,000 genetic loci.  Every transcript model in the index is represented on the microarray by at least one in situ-synthesized 60-mer oligonucleotide probe, for a total of 40,503 genes and 43,830 transcripts.  Dose-response linearity was evaluated, with  extremely high correlation (correlationmedian r = 0.98, with 50% of values between 0.99 and 0.93) between predicted and observed expression log ratios, and a compact distribution of regressions (median slope = 0.94, with 50% of values between 1.0 and 0.97).

While we have used similar microarrays to generate very reliable relative expression data in the past, we have not been able to estimate the absolute copy numbers of mouse transcripts, information which is critical to interpreting expression data, as well as to making decisions about follow-up studies of signature genes.  A set of synthetic “spike-in” RNA control transcripts and oligo probes, based on yeast intronic and intergenic sequences, allows us to estimate copy numbers of endogenous transcripts, as well as control for variations in target labeling and hybridization.

This platform was used to compare the expression profiles of embryonic stem (ES) cells and trophoblast stem (TS) cells.  This dData was compared to previous experiments, and providesproviding insight into what is thought to be the first differentiation step in mammalian development, - the transition from totipotent embryonic cells to multipotent trophoblast and inner cell mass.

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