Development of Vectors for the Regulated Expression of RNAi

摘要

The goal of this grant was to construct and test vectors that utilized the tetracycline repressor system and expression of short hairpin RNAs to regulate expression of genes in a conditional manner (plus or minus tetracycline) and to test the system in prostate cancer cell models to regulate genes of interest in prostate cancer. A series of vectors were constructed and tested for the regulation of Prostate Derived Factor (PDF) and the tumor suppressor p53. We did not see tetracycline-dependent inhibition of p53 expression. However, we found some LNCaP cell clones in which we saw up to 75% suppression of PDF expression. Further optimization of the vector did not generate clones with tighter regulation of expression. Thus, a retroviral vector directing tetracycline-regulated expression of a short hairpin RNA targeting specific genes was developed but proved only partially successful in achieving robust suppression of target genes in a tetracycline-dependent fashion.