The Interplay Between Estrogen and Replication Origins in Breast Cancer DNA Amplification
Annual summary rept. 1 Sep 2011-31 Aug 2012
BROWN UNIV PROVIDENCE RI
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In order to study the role that estrogen might have in breast cancer DNA amplification, I started to build the cell lines that I will use as a model for the upcoming experiments. MCF7 Flp-In cell lines have been established through the selection of cells stably transfected to carry an FRT Flippase Recombinase Target site sequence, which subsequently will function as acceptor site for the integration of constructs with an engineered replication origin via the FLPFRT method. The cell lines obtained were screened and characterized. Anticipating an experiment proposed for Year-Two, MCF7cMyc 6xERE cells, which have an ectopic c-Myc replication origin engineered to contain estrogen receptor-alpha binding elements, were treated with 17- estradiol for one month and subsequently exposed to a high dose of G418, whose resistance marker is encoded within the integrated pFRTc-Myc 6xERE construct. Although only one preliminary experiment has been done, the drug selection allowed enrichment of the cells that have DNA amplification at the ectopic c-Myc origin after cells were exposed to estrogen.
- Medicine and Medical Research