Molecular analysis reveals heterogeneity of mouse mammary tumors conditionally mutant for Brca1

Additional file 1:

Overall similarity of MMTV-driven tumor models. (A) Pearson correlation coefficient matrix for pairwise comparisons of log-intensity values among four spontaneous MMTV-PyMT tumors, based on 18,882 probes with reported values in at least 75% of samples (missing-value filter).(B) Pearson correlation coefficient matrix for pairwise comparisons of log-intensity values among four spontaneous MMTV-wnt1 tumors, based on 21,860 probes with reported values in at least 75% of samples (missing-value filter).

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Additional file 2:

Heterogeneity of Brca1 tumors. (A) H&E staining of original Brca1 tumors shows three representative types of morphology. A1 tumor shows features of adenosquamous carcinoma composed of sheets of squamous epithelial cells with eosinophilic cytoplasm and large leptochromatic nuclei. Some cells also form clusters of irregular glands. Several areas have whorls and clusters of spindle-shaped tumor cells suggestive of EMT. RP tumor shows features of, glandular carcinoma with no evidence of EMT. RA tumor has features of squamous nonkeratinizing Carcinoma. This tumor also does not show evidence of EMT. (B) Immunofluorescent detection of vimentin (red) and SMA (green) (right panels) shows mesenchymal features (vimentin) in tumor 0_A1. SMA stains fibroblasts only and not tumor cells, and is used here as control.

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Additional file 3:

List of most variable genes among mouse models with Brca1 deficiency. List of 339 probes that passed intensity and missing-value filters and had Standard Deviation (SD Gene Vector) >= 1.5 (to remove genes that have standard deviations of observed values less than 1.5, thus selecting for more variable genes across experiments), among 22 arrays encompassing the 5 original Brca1 tumors from our study, with 7 tumors from Brca1^+/-, p53^+/- irradiated (IR, Kohler) mice, and 10 tumors from Brca1^Co/Co, p53^+/-, MMTV-Cre (Furth) from a previous study [17]. Expression ratios for individual tumors are listed.

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Additional file 4:

List of most variable genes among original and passaged Brca1 mammary tumors. List of 416 probes that in addition to passing intensity and missing-value filters had a minimum 2.5-fold expression change in either direction from the median value in at least 20% of samples, encompassing 5 original independent Brca1 mammary tumors, 4 first-passage and 3 second-passage tumors. Expression ratios for original tumor 0_A1 and the average of expression ratios for transplanted tumors, as well as their fold differences are listed.

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Additional file 5:

Unsupervised Hierarchical Cluster. Unsupervised Hierarchical Cluster of genes and samples using 416 most variable genes across original and transplanted tumors (Additional file 8). Expression data from normal virgin mammary glands dissected from C57Bl6 and SCID mice were included in the analysis to define the contribution of genetic background to the differential gene expression. In the vertical axis, genes were clustered according to similarities in relative expression ratios. Gene clusters associated with genetic background are detailed on the side.

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Additional file 6:

Genotyping of Brca1 cell lines by PCR. Photographs of ethidium bromide-stained 1% agarose gels showing the products of PCR genotyping for p53 (A) and the Brca1^null allele (B) using genomic DNA from the indicated cell lines. L, molecular weight marker. C, p53 positive control. All cell lines used in this study were tested and showed loss of p53 allele and presence of the Brca1^null allele.

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Additional file 7:

Characterization of Brca1 cell lines by Western blot. Analysis of protein expression in whole cell lysates from 16 Brca1 cell lines using antibodies for Cytokeratin 5 (K5), Cytokeratin 18 (K18), c-kit (CD117), Vimentin, and Platelet-derived growth factor receptor alpha (Pdgfrα).

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Additional file 8:

Characterization of Brca1 cell lines by flow cytometry. Cell lines derived from anterior (A1.1 and A1.8) or posterior (P2.1 and P3.17) tumors were stained with fluorescently-conjugated antibodies against Cytokeratin 5 (A) or CD117 (B), in parallel with isotype control and analyzed by flow cytometry. In the histograms shown, the thick black line represents positive staining and the gray-filled lines represent the isotype control-matched mAb.

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