[초청강연] [Sigma-Aldrich Seminar] RNAi Strategies for Functional Genomics

- About the speaker: Dr. Heather Holemon is a principal investigator in the Functional Genomics group at Sigma-Aldrich. She obtained her Ph.D. from Washington University in Dr. Lee Ratner’s laboratory, where her thesis work focused on the function and packaging determinants of HIV-2 Vpx. After graduate school, she led Product Development groups, first at Incyte Genomics and then at ProteoPlex, Inc., in the development, validation and launch of an antibody-based microarray platform. After the acquisition of ProteoPlex, Inc. by EMD Biosciences in 2004, she took a principal scientist position at Orion Genomics where she worked on the development and validation of an assay used to detect epigenetic changes in cancer cells. - Abstract : (Subtittle : Gene silencing strategies for cancer research screening) RNA interference methodologies have greatly accelerated processes for dissecting and understanding gene function. Both synthetic siRNA and vector-based shRNA can be extremely useful for gene silencing experiments. The desired method depends on a variety of factors, including susceptibility to transfection and desired knockdown duration. We will discuss aspects of design, delivery, and in vivo gene silencing mediated by synthetic siRNAs. We will also discuss the delivery of short hairpin RNA (shRNA) using lentiviral vectors as a powerful means to mediate gene specific RNA interference in mammalian cells. Through the utilization of a recombinant lentivirus delivery system, the RNAi Consortium (TRC) library targets cell types that are often not amenable to synthetic siRNA transfection, and facilitates screens and experiments that require long-term, stable knockdown. We will discuss current validation efforts of this library and also demonstrate its applicability in different cell types. To further explore its utility in screening, we set out to develop strategies using the lentiviral-based shRNA libraries in larger scale silencing projects. Data presented here will demonstrate the use of shRNA-mediated knockdown in two separate cancer-screening studies. The first study used a tumor suppressor gene family set to screen for genes that could enhance cell sensitivity to the widely used cancer therapy drug, Paclitaxel. The second screen was performed to identify gene targets of a prostate cancer phenotype.