Generation of genetically tailored porcine liver cancer cells by CRISPR/Cas9 editing.

BioTechniques
Lobna ElkhadragyKyle M Schachtschneider

Abstract

Pigs provide a valuable large animal model for several diseases due to their similarity with humans in anatomy, physiology, genetics and drug metabolism. We recently generated a porcine model for TP53R167H and KRASG12D driven hepatocellular carcinoma (HCC) by autologous liver implantation. Here we describe a streamlined approach for developing genetically tailored porcine HCC cells by CRISPR/Cas9 gene editing and isolation of homogenous genetically validated cell clones. The combination of CRISPR/Cas9 editing of HCC cells described herein with the orthotopic HCC model enables development of various porcine HCC models, each with a specific mutational profile. This allows modeling the effect of different driver mutation combinations on tumor progression and in vivo testing of novel targeted therapeutic approaches in a clinically relevant large animal model.

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Methods Mentioned

BETA
transgenic
gene knockout
PCR
transfection
amplicon sequencing
FACS
ICE

Software Mentioned

CRISPOR
Fluidigm
ICE
Inference of Edits ( ICE

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