CRISPRi screening in cancer spheroids to investigate factors in genome stability

To investigate factors in genome stability in a 3D environment by performing a CRISPRi screen with cancer spheroids.

Project Overview

Although previous CRISPRi screening in 2D monolayers has provided useful knowledge on cancer drivers and therapeutic susceptibilities, it lacks an element of biological relevance to an in vivo environment. Therefore, our team was tasked with developing a cell culture method that is compatible with a 3D environment and CRISPRi screening in order to identify sources of DNA mutations in the tumor environment. On a high scale, the team must select a viable cell line for the screen, create and optimize a spheroid formation protocol, and develop a protocol to stain for γH2AX: a histone variant that acts as a sensitive marker for DNA damage.

Project Status

The team is continuing to passage cells, and are thawing new cells this week (week of 11/15). The team is finalizing the plate making, spheroid formation, and testing protocols as well as a materials list. The team has ordered materials and is waiting for them to arrive.