Overexpression of the Lycat transgene increased hematopoietic and endothelial cell gene expression in embryoid bodies. (A) Schematic representation of the Flk1:Lycat expression construct in which Lycat expression was driven by the mouse Flk1 promoter (Flk1P) and a PGK-neo cassette was inserted (top), and the vector control (VC) without Lycat cDNA (bottom). (B) Lycat mRNA was significantly increased in D4 EBs from transgenic ES clones of FC2, FC5, and FC22 as determined by semiquantitative RT-PCR. NC indicates negative control without input of DNA templates in the PCR reaction; β-actin, an internal RNA control. (C) The forced Lycat transgene increased hematopoietic and endothelial gene expression in D4 EBs from FC5 and FC22 ES clones detected by RT-PCR. Hematopoietic genes Gata1, Gata2, Runx1, CD41, Scl; endothelial genes Tie2 and Flk1; and β-actin as an internal RNA control. (D,E) The forced Lycat transgene increased mRNA expression of endothelial (panel D) and hematopoietic (panel E) but not endodermal (HNF3β; panel D) genes in D4 EBs from FC2 and FC5 ES clones detected by quantitative RT-PCR. Endothelial genes VE-Cadherin (Vecad), Flk1, CD31, Tie2; endodermal gene, HNF3β; hematopoietic genes Scl, Gata1, Gata2, Runx1, CD41, β-H1 hemoglobin (β-H1), and β-major hemoglobin (β-Major). The mRNA levels were normalized by GAPDH. (D,E) Error bars represent standard deviations.