I met with a problem that when infected with retrovirus, my 3T3-L1 cells cannot be differentiated and died a few days after differentiation medium induction. Before infection, 3T3-L1 cells grew healthy and can be differentiated.
I started differentiation 2 days post 100% confluence, used differentiation medium containing 5ug/ml insulin, 1uM DEX and 0.5mM IBMX for the first 2 days, shifted to medium containing 5ug/ml insulin in the following days and refreshed medium every 2 days. Cells started to die about 3-5 days after induction and no differentiated cells could be found.
I used pLkO.1 retrovirus vector for siRNA knock-down.
I tried different virus titers but got the same result.
Would you please help me with this problem? Many thanks.