Cloning and Characterization of Active Egr-1 Target Genes by In Vivo Crosslinking
Abstract
The purpose of this project is to identify the profile of Egr-1 target genes in breast cells. Normal breast cells are shown to express Egr-1, while breast cancer cells do not. It is, therefore, important to identify the nature of those target genes regulated by Egr-1 which are absent in breast cancer cells. I have approached this goal by performing in vivo crosslinking of Egr-1 to its target sites in breast cells, followed by immunocapture of Egr-1 together with its targets. In this report, I have confirmed expression of Egr-1 in normal, but not in breast cancer cells. Furthermore, I have succeeded in capturing Egr-1 and its target DNA sites by immunoprecipitation. In addition, I have proceeded by cloning the Egr-1 target DNAs for further characterization. Significantly, I have also developed a technique of multiplex PCR using the captured DNA as primers to identify those Egr-1 binding sites adjacent to, and possibly regulating, genes from a cDNA library. This technique will be used to focus on Egr-1 targets which affect gene expression.
Document Details
- Document Type
- Technical Report
- Publication Date
- May 01, 2000
- Accession Number
- ADA382458
Entities
People
- Ian De Belle
Organizations
- Sanford Burnham Prebys Medical Discovery Institute