Tyronsine Kinase Gene Expression Profiling in Prostate Cancer
Final rept. 1 Mar 2000-28 Feb 2004
CALIFORNIA UNIV BERKELEY LAWRENCE BERKELEY LAB
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The expression of protein kinases genes if often altered in tumors. The aberrant expression of several of these genes typically parallels the progression toward a more malignant phenotype. We developed a cDNA micro-array-based screening system to mearuser the level of expression of tyronsine kinase tk genes. The hardware for preparation of cDNA micro-arrays and basic protocols for hybridization were developed in year 1. In the second year, we finished cDNA synthesis from prostate cancer cell lines and 6 frozen tissue specimens. We continued our DNA sequencing effort and added additional targets to our micro-arrays. Using prostate cancer cell lines, the system delivered reproducible results about tk gene expression during cell transformation and progression toward a more malignant phenotype. Comparing the absolute expression levels form cDNA micro-arrays with data from Northern blot analyses suggested that our initial approach using mixed-based oligonucleotide primers led to lowered representation of highly abundant transcript. This problem has been addressed with a new primer design to be better suited to investigate the tk gene expression in small samples. We then applied our protocols to the gene expression analysis of micro-dissected tissues, and found reliable detection of abl gene overexpression with an RNA equivalent of 3 cells.
- Anatomy and Physiology
- Medicine and Medical Research