CRISPER Cas9 Mediated Knock - In of Fluorescent tag to track GATA6 in Mouse Embryonic Stem Cells

Abstract

The embryonic stem cells (ESC) are pluripotent cells that are derived from the inner cell mass of the blastocyst and have the ability to differentiate to almost all cell types, in-vitro and in-vivo. Cells respond to the extracellular environment with a set of receptors which is altogether known as mechanical signaling. Recent studies reveal that the mechanical environment affect the behaviour and function of different types of cells, including stem cells. However, signaling pathways involved in the mechanical regulation of stem cell properties remain largely unknown. During early mammalian development, once the inner cell mass is formed, next cell fate choice is formation of Epiblast and Primitive Endoderm (PE). Nanog is an epiblast marker and marker for PE is GATA-6. In this study, an endogenous GATA-6 BFP ESC line is created by CRISPR-CAS9 mediated Knock-In. This reporter cell line will be used to detect/track primitive endoderm cells during ES cell differentiation in EBs and blastocyst.