Using RNA Interference to Reveal Genetic: Vulnerabilities in Human Cancer Cells
Abstract
A major barrier to understanding breast cancer is the lack of comprehensive and systematic large scale studies that provide functional information about the entire genome. These insights can be obtained through RNAi (RNA interference) genetic studies RNAi is a cellular process that regulates gene expression in a sequence specific manner. We have developed a library of plasmids expressing shRNAs that engage the endogenous RNAi pathway and produce mature siRNAs that efficiently target any gene of interest. We have generated more than 200,000 constructs that allow us to perform loss of function studies of almost every gene in the human genome. Furthermore, we have developed a microarray-based analytical platform that facilitates the study of thousands of genes concurrently in pools. We conducted a screen to detect resistance to anoikis (cell death trigged by loss of attachment to the extracellular matrix, ECM) in the MCF10A breast epithelial cell line. Our screen of 1,500 shRNAs resulted in identifying the well known tumor suppressor, Pten, as an attenuator of anoikis among other candidate genes. In addition, we have validated an in vitro anoikis assay as an approach to identify putative tumor suppressors involved in breast epithelial cell transformation.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jul 01, 2006
- Accession Number
- ADA460501
Entities
People
- Despina Siolas
- Gregory Hannon
Organizations
- Cold Spring Harbor Laboratory