a) Your lab has produced a mouse which is a knockout for the LNO gene. This gene is a known receptor for the SVU growth factor. This is an important receptor during development and loss of this gene causes embryonic lethality in your mouse. The embryos do survive for 14 days in utero. In order to study the loss of this gene, you decide to create a cell line from the mouse embryos. How would you do this? b) You want to determine the specific binding of the SVU to the LNO receptor to find the Kd of the receptor and the number of receptors per cell. How would you do this? What would be your controls? c) Surprisingly, you get similar binding curves for cell lines expressing the LNO receptor and the cell line from your knockout mouse. You are worried your mouse was not really a protein null! How could you test for presence of the LNO receptor in the knockout cell line? d) Results from the previous experiment show that there is no expression of the LNO receptor in your cell line and you conclude there must be another receptor that binds the SVU ligand. How could you identify this receptor? e) You find one novel receptor LOCI also binds the SVU growth factor. You want to perform a binding assay but do not have a similar cell line which does not express this receptor. How would you perform this binding assay?
