CRISPRi screening in cancer spheroids to investigate factors in genome stability

Develop a cell culture method compatible with a 3D environment and CRISPRi screening in order to identify sources of DNA mutations in the tumor environment through γH2AX expression.

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 finishing up the preliminary report, planning spheroid experiment 3, planning qPCR timeline, and will be completing Passage 12-15 this week.