Genetic Studies of Natural Glycosphingolipid Ligands for NKT Cells

Glycosphingolipids (GSL) are natural ligands of NKT cells. Several laboratories have reported the in vitro activity of isoglobotriosylceramide (iGb3) in stimulating NKT cells. However, the knockout mice of iGb3 synthase showed no deficiency in development and function of NKT cells. There is a lack of knowledge on the genetics of redundant natural glycosphingolipid ligands. We have identified additional glycosphingolipid with stimulatory activity to NKT cells, including fucosyl lactosylceramide (H antigen). Here we describe the procedures to generate mice with deficiencies in Fut1, Fut2, and Sec1 genes to deplete H antigen through BAC engineering for the generation of ES cell-targeting construct, as well as the mice with deficiency of both blood group H-GSL ligand and isoglobotriosylceramide.