Hormonal Involvement in Breast Cancer Gene Amplification
Final rept. 24 Sep 2007-23 Sep 2010
BROWN UNIV PROVIDENCE RI
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Our specific aims were to map origins of replication in the breast cancer genome and compare these sites with genomic positions that bind the estrogen receptor ER and genomic regions of DNA amplification. Correlations would support our hypothesis that ER adjacent to replication origins may interact with the replication machinery to drive DNA amplification, hallmark of many cancers. We began our experiments with the well-studied breast cancer cell line MCF7. We refined the lambda exonuclease method to obtain nascent DNA strands up to 20 times enriched and demonstrated that the myc origin maps to the same position in HeLa and MCF7 cells. We size fractionated nascent DNA 500-1500 nt for next generation sequencing which gives better resolution and sensitivity than DNA microarrays. The results of Illumina sequencing revealed that our nascent DNA sample from the whole genome included many of the origins previously reported for 1 of the human genome. Development of this technology serve as the basis for our IDEA Extension grant which was funded to extend the work begun in this parent grant.
- Anatomy and Physiology
- Medicine and Medical Research