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Searched keyword : CRISPR
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CRISPR-Cas9 tools have recently emerged as a very powerful way to edit genomes and control gene expression. The Cas9 protein is an endonuclease that can be guided by a small guide RNA (gRNA) to any target DNA sequence of choice. This feature has enabled the construction of gRNA libraries to perform genome-wide CRISPR-Cas9 functional screens. Such screens have notably been used to identify factors involved in drug and toxin resistance in human cells. We are now in the process of developing such a screen for bacteria. In this screen, we use the catalytic null mutant of Cas9 known as dCas9 to block transcription at target positions and knockdown genes. The purpose of this project is to investigate the properties of this type of screen in bacteria.
We have recently identified around 500 long non-coding RNAs that follow a very precise expression pattern in undifferentiated mouse Embryonic Stem cells. In order to assess if any of those molecules are functionally relevant for the biology of ES cells we will perform a functional screening based on the CRISPR technology. We plan to use a modified version of the CRISPR-CAS9 technology, the CRISPRon system, in which (i) the guide-RNAs (gRNAs) will target the promoters of our candidate lncRNAs (at least 5 gRNAs per promoter) and (ii) an enzymatically-inactive CAS9 will be engineered to recrute 10 molecules of the potent VP64 transactivator. This will enable to upregulate the targeted lncRNA from its endogeneous locus.
CRISPR polymorphism is a powerful tool to subtype Salmonella strains and is now used in routine for epidemiological investigations. The aim of this project is to transfer and upgrade a published and worldwide used webtool to extract the spacer content from fasta sequences or paired-end reads.