In recent years, we have worked towards extending the E-MAP technology to functionally interrogate mammalian biology. To this end, we developed a combinatorial RNA interference (RNAi) based platform for the systematic and quantitative generation of genetic and chemical-genetic interaction maps in mammalian cells. The approach relies on high-throughput microscopy and automated liquid handling to minimize time, labor and human error. The microscopy-based nature of our platform allows us to quantify not only proliferation, but also other phenotypes that can be identified via cell staining or reporter cassettes. By combining chemicals with RNAi knockdowns, we can further interrogate the interplay between cellular machinery and drug treatments. This feature also makes our platform an effective tool for identifying genetic backgrounds that are sensitive to particular drugs.
Most recently we have worked with the Qi lab at Stanford and Mali lab at UCSD to develop an experimental methodology for generating quantitative genetic interaction maps in mammalian cells utilizing CRISPR and CRISPRi/a. In a close collaboration with the Qi lab we developed a platform for performing pooled CRISPRi genetic interaction screens and applied it to a set of over 100 chromatin-related factors in a human cell culture model (HEK293). Our approach utilizes a stable expression of an inducible dCas9-KRAB construct combined with a library of single or double sgRNA (single guide RNA) constructs delivered via lentiviral transduction. Relative abundance of individual sgRNA combinations is determined through next generation sequencing. In collaboration with the Mali lab at UCSD we developed a similar approach utilizing the CRISPR gene editing toolkit and applied it to a set of tumor suppressor genes and druggable targets in two different human cell lines (HeLa and A549). We have also developed a suite of new software tools aiding the processing and scoring of the raw data that result from these pooled screens.