Additional file 1:

qPCR analysis of Krt14, Krt18 and Esr1 expression in mammary epithelial subpopulations. The data describes qPCR analysis of expression of Krt14, Krt18 and Esr1 in triplicate independent samples of CD24^+/Low Sca-1^- cells, CD24^+/High Sca-1^- cells and CD24^+/High Sca-1⁺ cells. Each data point is the mean level of expression, ± 95% confidence intervals, across the three samples of that population relative to the comparator sample. A 'round robin' comparison was used as described in the Methods. Genes considered to be characteristic of the comparator population are indicated next to each pair of graphs. Krt14 expression was undetectable in the CD24^+/High Sca-1^- and CD24^+/High Sca-1⁺ cells. Krt18 expression was undetectable in the CD24^+/Low Sca-1^- cells.

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Additional file 2:

Relative expression levels of 2182 Affymetrix probes across three virgin mouse mammary epithelial subpopulations. The spreadsheet gives the relative expression levels for all differentially expressed probes across all three populations. Expression levels are indicated by a relative abundance score for each populations. A high positive value indicates expression at a high level, a low negative score indicates very low expression levels. The Affymetrix probe ID, Gene Symbol and q-value (indicating the % false discovery rate) are also indicated.

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Additional file 3:

Full heat map and hierarchical cluster analysis of differential gene expression across virgin mammary epithelial populations. The image shows heat map clustering of differentially expressed genes across the three cell populations. Red indicates high expression, green indicates low expression.

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Additional file 4:

Genes characteristic of basal/myoepithelial cells. The table shows all 861 genes in the basal/myoepithelial population with an abundance score of 2 or more when the 1427 differentially expressed gene set was sorted by descending abundance scores in the basal/myoepithelial population. Such genes were considered characteristic of the population. Where differential gene expression was indicated by more than one probe, an average value for each of the contrasts across the probes was calculated. The number of probes is indicated.

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Additional file 5:

Genes characteristic of luminal ER^- cells. The table shows all 326 genes in the luminal ER^- population with an abundance score of 2 or more when the 1427 differentially expressed gene set was sorted by descending abundance scores in the luminal ER^- population. Such genes were considered characteristic of the population. Where differential gene expression was indicated by more than one probe, an average value for each of the contrasts across the probes was calculated. The number of probes is indicated.

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Additional file 6:

Genes characteristic of luminal ER⁺ cells. The table shows all 488 genes in the luminal ER⁺ population with an abundance score of 2 or more when the 1427 differentially expressed gene set was sorted by descending abundance scores in the luminal ER⁺ population. Such genes were considered characteristic of the population. Where differential gene expression was indicated by more than one probe, an average value for each of the contrasts across the probes was calculated. The number of probes is indicated.

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Additional file 7:

Summarised gene expression microarray and qPCR gene expression analysis for 58 test genes. The table shows a comparison between the gene expression patterns determined by microarray analysis and those determined by qPCR. The gene expression microarray relative abundance scores are summarised as follows: --- = -32 to -22, -- = -22 to -12, - = -12 to -2, +/- = -2 to +2, + = 2 to 12, ++ = 12 to 22, +++ = 22 to 32, ++++ = 32 to 42. Where more than one identifier for a gene scored as differentially expressed, the mean score across all the identifiers was used to determine the summarised microarray score. The summarised score was in turn used to define the array-based expression pattern, with a score of +, ++, +++ or ++++ indicating that a gene was expressed in a particular population. In some cases, the genes were expressed in more than one population. The summarised qPCR expression pattern is based upon the patterns of gene expression determined from Figure 3. Whether or not the array and qPCR based analyses show concordance in their assignment of expression patterns is indicated. NDE = no differential expression in microarray. N/A = comparison cannot be made as qPCR probe failed. *In these comparisons, technical failure of the Affymetrix probe cannot ruled out.

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Additional file 8:

Comparison of numbers of genes identified in common between basal/myoepithelial, luminal ER^- and luminal ER⁺ cells and previously published datasets. The table compares previously published datasets with the subpopulation specific genes identified in the current analysis. Published lists of genes [27,28], probes [6] or SAGE tags [29] significantly differentially expressed between mouse basal mammary stem cell enriched/myoepithelial cells compared to in vitro colony forming cells (luminal cells) [6], human CD10^- CD44⁺ basal cells compared to CD24⁺ luminal cells [29] or human CD10⁺ myoepithelial cells compared to EMA⁺ luminal cells [27,28] were condensed to remove multiple probes or tags against the same gene and to identify only well-annotated genes. The distribution of the differentially expressed genes in the basal/myoepithelial, luminal ER^- and luminal ER⁺ gene lists from the current study was then determined.

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Additional file 9:

List of genes common to basal/myoepithelial cells and previously published basal or myoepithelial datasets. The table lists those basal/myoepithelial genes found in previously published datasets which were also found the basal/myoepithelial population in the current study.

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Additional file 10:

List of genes common to luminal cell subpopulations and previously published luminal datasets. The table lists those luminal genes found in previously published datasets which were also found the luminal populations in the current study.

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Additional file 11:

Network interaction map for basal/myoepithelial specific genes. Interaction data between basal/myoepithelial specific genes based on physical interactions (black lines) and interactions in complexes (brown lines) with no interpolated genes used. The nodes are colour coded to indicate relative strengths of expression of the gene within the cell population. Brighter reds indicate highest levels of expression. Darker reds indicate genes less strongly expressed (although still with enriched expression within the population compared to the other cell types).

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Additional file 12:

Network interaction map for luminal ER^- specific genes. Interaction data for luminal ER^- specific genes based on physical interactions (solid lines) and transcriptional interactions (dashed lines). The nodes are colour coded to indicate relative strengths of expression of the gene within the cell population. Brighter reds indicate highest levels of expression. Darker reds indicate genes less strongly expressed (although still with enriched expression within the population compared to the other cell types). White nodes indicate interpolated genes used by the network mapping software to extend and link the network.

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Additional file 13:

Network interaction map for luminal ER⁺ specific genes. Interaction data for luminal ER⁺ specific genes based on physical interactions (solid lines) and transcriptional interactions (dashed lines). The nodes are colour coded to indicate relative strengths of expression of the gene within the cell population. Brighter reds indicate highest levels of expression. Darker reds indicate genes less strongly expressed (although still with enriched expression within the population compared to the other cell types). White nodes indicate interpolated genes used by the network mapping software to extend and link the network.

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Additional file 14:

Identification of prominent modules of differentially expressed genes in the luminal ER^- and luminal ER⁺ networks. The results of the first, second and third pass analyses for network modules in the luminal ER^- and luminal ER⁺ networks are shown. Rectangular nodes are first pass nodes, octagonal nodes are second pass nodes and small green oval nodes are third pass nodes. Thick red lines are first pass connections, medium size red lines are second pass connections and thin red lines are third pass connections. Black rectangles indicate differentially expressed hubs for which no modules could be built. Coloured rectangles indicate module groupings of differentially expressed genes. Solid lines indicate physical interactions, dotted lines transcriptional interactions.

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Additional file 15:

Topology of interaction modules within the luminal ER^- network. Luminal ER^- interaction modules are shown projected on to the luminal ER^- network. First, second and third pass nodes are indicated as coloured rectangular, octagonal and oval nodes respectively. First, second and third pass connections are indicated as thick, medium and thin red lines respectively. Solid lines indicate physical interactions, dotted lines transcriptional interactions. The different colourings of the first and second pass nodes indicate module groupings of differentially expressed genes. Third pass nodes are coloured green.

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Additional file 16:

Topology of interaction modules within the luminal ER⁺ network. Luminal ER⁺ interaction modules are shown projected on to the luminal ER⁺ network. First, second and third pass nodes are indicated as coloured rectangular, octagonal and oval nodes respectively. First, second and third pass connections are indicated as thick, medium and thin red lines respectively. Solid lines indicate physical interactions, dotted lines transcriptional interactions. The different colourings of the first and second pass nodes indicate module groupings of differentially expressed genes. Third pass nodes are coloured green. Black rectangles indicate differentially expressed hubs for which no modules could be built.

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Additional file 17:

Network metrics for the luminal ER^- and luminal ER⁺ module subnetworks. The table gives the values for the parameters describing the luminal ER^- and luminal ER⁺ networks both with and without the identified modules. Network manipulations were performed in Cytoscape [111], and the average network clustering <C>, average connectivity <k>, power exponent γ and mean shortest path <l> were derived with the Cytoscape Random Networks plug-in. Note that due to the highly fragmented nature of the non-module subnetwork, the mean shortest path does not constitute a reliable metric.

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Additional file 18:

List of genes common to mammary cell subpopulations and previously published datasets of estrogen-responsive genes. The table lists estrogen-responsive genes identified in previously published datasets which were also found in the mammary epithelial cell subpopulations in the current study.

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Additional file 19:

Genes with potential roles in lineage selection/cell fate determination through paracrine signalling or as transcriptional regulators. The table lists those genes from the three populations whose protein products have potential roles in intercellular signalling or as transcriptional regulators. *Distribution confirmed by qPCR.

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Additional file 20:

Sox6 over-expression maintains luminal differentiation in mammary epithelial cells in vitro. Additional images of immunofluorescence staining for keratin 14 (A, B) and keratin 18 (C, D) expression in primary mouse mammary epithelial cells transduced with lentivirus expressing Sox6 and GFP. A, C, bars = 30 μm. B, D, bars = 60 μm. Arrows in A indicate rare Sox6-GFP cells which are also weakly K14 positive. The majority of Sox6-GFP cells are K14 negative.

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Additional file 21:

Genes examined by qPCR analysis. The table gives the gene name, symbol, TAQMAN Assays on Demand (Applied Biosystems) assay reference and Unigene ID for all qPCR probes used.

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