"Live-cell imaging combined with kinetic analyses has provided new biological insights on the gene expression pathway. However, such studies in mammalian cells typically require use of exogenous over-expressed gene constructs, which often form large tandem gene arrays and usually lack the complete endogenous regulatory sequences. It is therefore imperative to design methodology for analyzing gene expression kinetics of single alleles of endogenous genes. While certain steps have been taken in this direction, there are many experimental obstacles standing in the way of a robust genome-wide system for the in vivo examination of endogenous gene expression within the natural nuclear environment. GENEXP sets out to provide such a system.
It will start with methodology for robust tagging of a multitude of endogenous genes and their transcribed mRNAs in human cells using the ""CD tagging"" approach. Thereby, in vivo mRNA synthesis at the nuclear site of RNA birth will be explored in a unique manner. A high-resolution study of gene expression, in particular mRNA transcription and mRNA export, under endogenous cellular context and using a genome-wide live-cell approach will be performed. GENEXP will specifically focus on the:
i) Transcriptional kinetics of endogenous genes in single cells and cell populations;
ii) Kinetics of mRNA export on the single molecule level;
iii) Examination of the protein composition of endogenous mRNPs;
iv) High throughput scan for drugs that affect gene expression and mRNA export.
Altogether, GENEXP will provide breakthrough capability in kinetically quantifying the gene expression pathway of a large variety of endogenous genes, and the ability to examine the generated molecules on the single-molecule level. This will be done within their normal genomic and biological environment, at the single-allele level."
Field of science
- /social sciences/sociology/demography/fertility
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