GALK1 Stable Knockout HEK293T Cell Line

The Galactokinase 1 (GALK1) is major enzyme in the Leloir pathway, the primary pathway for galactose metabolism. Specifically, this enzyme modifies galactose with a phosphate group to generate galactose 1-phosphate which serves as a precursor for O-linked glycosylation. Deficiencies in GALK1 leads to Galactosemia: a congenital condition that causes galactose to accumulate in blood and tissue. GALK1 Stable Knockout cell line was generated through the knockout of the GALK1 gene in HEK293T cells using the CRISPR-Cas 9 system. Cells were transiently transfected with an expression vector containing a GFP tag and a Guide RNA sequence that targeted GALK1. Clones were initially chosen through strong GFP expression and knockout of GALK1 confirmed via sequencing and GALK1 enzymatic activity. In combination with the GALE GALK1 Stable Knockout cell line (T6151), this line may prove useful in the study of O-linked glycosylation and galactose metabolism.