2014.9.22 RNA-guided Genome Engineering Using CRISPR-Cas System: Technology and Application
题目：RNA-guided Genome Engineering Using CRISPR-Cas System: Technology and Application
报告人：Dr. Le Cong
Broad Institute of MIT and Harvard, Massachusetts Institute of Technology, 75 Ames Street, Cambridge, MA 02142, USA.
The integration of genetic, biochemical, and engineering techniques with vast amount of sequencing data have enabled us to decode the genetic information of different biological systems with unprecedented resolution. However, this process has been limited by the lack of powerful and precise tools to control biological systems at genome-scale, especially in eukaryotes. Genome engineering technology enables the targeted modification and modulation of genome sequences, with wide applications in basic research and medicine. One of the most exciting recent advances in this field is the development of RNA-guided designer nuclease based on Type II clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system. We and many other groups have previously shown the effectiveness, robustness, and scalability of this technology in editing mammalian genomes in vitro. Beyond these studies, further improvement of its efficiency and specificity, and the in vivo delivery of this system in animal models are key steps towards further applications of the CRISPR-Cas9 system.
Here we present our recent progress on the improvement of the CRISPR-Cas9 technology, particularly in vivo editing of target genomic loci in adult animals using a novel CRISPR-Cas9 system delivered by adeno-associated virus (AAV). We developed this new orthologous Cas9 to allow the inclusion of all required components of CRISPR-Cas9 system in a single AAV vector. To demonstrate the functionality of our design and establish its therapeutic potential, we chose to use this system to edit genes in mouse liver whose homologous human versions are associated with metabolic diseases. We also characterized its efficiency and specificity compared with the widely used S. pyogenes Cas9 system. Our results exhibit the expandability of the CRISPR-Cas9 tools, the benefit of developing orthologous Cas9 system, and the applicability of CRISPR-Cas9 system for in vivo genome engineering.
Overall, RNA-guided genome engineering systems will lead to transformative paradigms in basic research, industrial-scale bioengineering, disease modeling, and gene therapy. It will greatly enhance our ability to translate biomedical research into clinical application, enabling breakthroughs in the study and treatment of many refractory human genetic disorders, such as metabolic diseases, cancer, and neuropsychiatric diseases.