Genome Engineering Technology and Its Application in Mammalian Cells.

摘要

The advancement of high-throughput, large-scale biochemical, biophysical, and genetic technologies has enabled the generation of massive amounts of biological data and allowed us to synthesize various types of biomaterial for engineering purposes. This enabled improved observational methodologies for us to navigate and locate, with unprecedented resolution, the potential factors and connections that may contribute to biological and biomedical processes. Nonetheless, it leaves us with the increasing demand to validate these observations to elucidate the actual causal mechanisms in biology and medicine. Due to the lack of powerful and precise tools to manipulate biological systems in mammalian cells, these efforts have not been able to progress at an adequate pace.;This work aims to bridge the gap between data generation and experimental verification by developing molecular-resolution genome engineering technologies that can effect cell-specific genetic and epigenetic perturbation in mammalian cells. The development of these novel tools starts from harnessing aspects of two prominent families of microbial systems, the Transcription Activator-like Effectors (TALEs) and the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas systems. The development of these technologies was executed through several lines of engineering efforts. When applied in higher eukaryotes, these tools provide researchers with new reverse engineering instruments to directly probe relevant biological molecules and pathways that are observed from analysis of biological data. In combination with sensitive and accurate readout methods in mammalian systems, these technologies could together establish transformative means for modeling the causal relationships between genetic or epigenetic variances and human disease, while also serving as the first step towards the development of rational molecular therapies for complex human disorders and of synthetic biology applications for the research community.