Journal: PLoS ONE

Article Title: MicroRNA-27a Contributes to Rhabdomyosarcoma Cell Proliferation by Suppressing RARA and RXRA

doi: 10.1371/journal.pone.0125171

Figure Lengend Snippet: A ) Experimental design: the miRNA population from each cell line was compared to a common reference sample consisting of a balanced mixture of eight small RNA samples (< 200 nt) prepared from the same cell lines. Two replicates of each experiment were performed using different microarray slides in which sample and reference RNAs, labeled with Cy3 (gray arrow) or Cy5 (black arrow) fluorochromes, were crossed in both combinations (dye-swapping procedure). B ) Heat map of 16 discriminant miRNAs between PAX3/FOXO1 positive ARMS (in gray on the right, RH4, RH30, RH28) and negative (in dark grey on the left, RD, SMS-CTR, RH18, RH36, CCA) RMS cell lines identified by SAM analysis ( rows : miRNAs; columns : RMS cell lines). A color-coded scale for the normalized expression values was used as follows: yellow and blue represent high and low expression levels in RMS cell lines with respect to a reference sample (a pool of eight RMS cell lines). The expression level of each miRNA was calculated as the ln (RMS cell line/Pool). C ) Expression levels of miR-199a, miR-23a and miR-27a in eight RMS cell lines obtained with qRT-PCR. Two independent experiments were performed in triplicate. Results are shown as relative expression ratio obtained with the 2 -ΔΔCt method. RNU6B was used as reference miRNA. Vertical bars represent the 95% confidence interval (IC).

Article Snippet: Array scanning was carried out using a GSI Lumonics LITE dual confocal laser scanner with a ScanArray Microarray Analysis System (Perkin Elmer, Waltham, MA), and raw images were analyzed with QuantArray Analysis Software (GSI Lumonics, Ottawa, Canada).

Techniques: Microarray, Labeling, Expressing, Quantitative RT-PCR