Validating transcripts with probes and imaging technology

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Materials Processing Center; Massachusetts Institute of Technology. Koch Institute for Integrative Cancer Research at MITHigh-throughput gene expression screens provide a quantitative picture of the average expression signature of biological samples.However, the analysis of spatial gene expression patterns with single-cell resolution requires quantitative in situ measurement techniques.These cells can then be used for either bulk measurements or for single cell measurements that require cell lysis, such as quantitative RT-PCR.Similarly, sorting GFP positive cells from transgenic mice expressing GFP under the control of a cell-type specific promoter of interest enables enriching for the cells expressing this gene and allows characterization of the expression program of these particular cells, as has been done with tissue stem cells.High-throughput gene expression screens provide a quantitative picture of the average expression signature of biological samples.High throughput gene expression screens provide a quantitative picture of the average expression signature of biological samples.

These methods, which are based on advances in probe design, imaging technology, and image processing, enable the absolute measurement of transcript abundance in individual cells with single-molecule resolution.Additionally, cancer cells often show profound diversity not only in their transcript content but also in their genotype.This diversity stems from increased mutation rates, rapid cell proliferation and spatially varying selection forces.The precise location of cells within the tissue translates to constant changes in the levels of niche-secreted morphogens, which give rise to position-modulated gene expression programs.Thus two adjacent cells could harbor dramatically different expression programs.

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