Figure 4

Sense and antisense expression profiles. (a) Unsupervised cluster analysis of expression profiles detected by C (blue), N (red) and CN (grey) capture probes across 9 cell lines. A: UACC812; B: ZR75-30; C: BT474; D: MDA361; E MCF10A, F: SKBR3; G: UACC893; H: MDA453 and I: CAMA1. Columns in the gene expression matrix correspond to IMAGE clone IDs. Each cell line is represented by one row. Minimum presence per cDNA clone was set to 7 out of 9 in each data set. (b) Correlation between N- and C-probes across hybridizations A to I for 430 probe pairs, i.e. IMAGE clones. Frequency of Pearson correlation is shown as bars with a bin width of 0.05 (gray bars). For each frequency, the contribution of probe pairs from IMAGE clones previously reported as antisense transcribed is shown as striped areas. Correlation calculations are performed on log2ratios using within hybridization mean log2ratio per reporter across features with SNR > = 10 in either channel, i.e. Cy3 or Cy5. IMAGE clones were removed if their respective N- and C- probes were suspected to potentially cross-hybridize or if their number of missing values across hybridizations was > 2 for either N- or C- probe. (c) Schematic representation of the overlap between KHK (BC006233) and CR623121 transcripts and rational of the RT-PCR reaction aimed to verify the observed coordinated regulation of sense and antisense transcripts. Black arrows represent orientation of transcription for each RNA. Grey arrows mark positions for the PCR primers used in (d). Distance between primers positions refer to the final amplification products. (d) Relative expression of KHK and its antisense CR623121 detected by RT-PCR across all cell lines. Two amplicons of 406 and 626 bp corresponding to KHK sense and antisense transcripts were obtained. Lower panel: ethidium bromide staining after separation in 1% agarose. Band intensities were quantified with the Kodak 1D Image Analysis Software and mean intensities were plotted.