Year of Award:
Molecular & Cellular Analysis Technologies
Other PI or Project Leader:
NORTH CAROLINA STATE UNIVERSITY RALEIGH
Project SummaryIt has been clear for over a decade that cancer is not a single disease and that this heterogeneity is a primarybarrier to the understanding of oncogenic mechanisms and treatments. Cancers vary epigenetically andgenetically at multiple length and time scales and between patients. There can be many distinct mechanismsdriving oncogenic and metastatic processes, even within a single cancerous cell. The challenge researchers andclinicians face is how to understand cancer from the perspective of simultaneous perturbations to multiplegenes and proteins. Risk variants identified through genome wide association studies and epiGWAS can beindividually perturbed in large experiments comprised of many 384-well plates through RNAi and CRISPRlibraries, and even be combinatorially perturbed and screened in `one-pot' using barcoding strategies.However, even state-of-the-art CRISPR screening methods are restricted to functional perturbations of 2 or 3variants at a time per cell. These restrictions arise from the difficulties repetitive sequences in gRNA arrayspresent in both expression construct synthesis and stability. Here we propose a new method that will becapable of expressing randomized combinatorial libraries of thousands of distinct gRNAs, with each cell of apopulation expressing an array of over 30 gRNAs.