Expression data for validation of single cell cDNA amplification method (V1V3 method)

A systems-level understanding of a small but essential population of cells in development or adulthood (e.g., somatic stem cells) requires accurate quantitative monitoring of genome-wide gene expression, ideally from single cells. We report here a strategy to globally amplify mRNAs from single cells for highly quantitative high-density oligonucleotide microarray analysis that combines a small number of directional PCR cycles with subsequent linear amplification. Using this strategy, both the representation of gene expression profiles and reproducibility between individual experiments are unambiguously improved from the original method, along with high coverage and accuracy. Keywords: Method validation To verify the ability of the developed method, we prepared total RNA from ES cells and diluted it to the single-cell level (10 pg). Eight array data from the cDNA samples amplified independently from the 10 pg RNA (amplified samples) were compared to each other, and to eight array data from the undiluted 5 ug RNA (nonamplified controls). Since all the materials were the same, all of the observed variations were attributable to the methodogoly.