- CORE B The Nanotechnology-based Transcription Core B will provide support for gene expression analysis and gene knock-down to all three projects of the PPG. We have been successful in our effort to develop a multiplex gene expression profiling resource at our center. In collaboration with Aviv Regev at the Broad Institute of Genetics and Genomics, we have developed technologies to undertake multiplex gene expression profiling using newly acquired NanoString nCounter technology and developed analytical tools to analyze the data obtained. With the support from the Broad Institute, our expression Core has the know-how, technology, resources, and analytical skills to support the research projects of this PPG. The Core allows for multiplex gene expression analysis of up to 500 genes in a single RNA sample using a recently acquired NanoString nCounter, which saves both time and material. Indeed, NanoString nCounter technology is ideally suited for gene expression analysis in rare cell populations such as those obtained from tumor or CNS tissue and thus is perfectly suited for the projects in the PPG. In addition to multiplex gene expression analysis, the core will also provide technology and support for knock- down of specific genes in primary T cells using silicone nanowire (NW)-based technology. Since current methods for gene knock-down are not amenable for use in primary exhausted T cells, Dr. Hongkun Park at Harvard University will participate in the Core as the Co-Investigator and provide vertical silicone nanowires and know-how to each of the projects. The silicone nanowires will be coated with siRNA to knock-down specific genes in exhausted T cells. The effect of specific gene knock-down on cell phenotype and function will be undertaken in the individual projects, but primary data on gene expression following gene knock-down will be generated by the Core. The core has been set up so that gene expression data from various cell types and following siRNA nano-wire knock-down obtained from different projects can be readily analyzed and compared. The data will be archived, analyzed, and stored in a designated drop-box folder such that all the investigators of the PPG will have access to the expression and knock-down data. Furthermore, the data will be discussed monthly by video/teleconferencing.
- CORE B The multiplex gene expression and gene 'knock-down' resources of the core will allow us to develop a molecular map to understand the development, function, and regulation of TIM-3 expression in both mouse and human T cells in different disease settings. Identification of the transcriptional mechanisms that regulate TIM-3 expression could be used for developing therapies for multiple sclerosis, cancer, and chronic microbial infections.
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