I am in the process of attempting to immortalize some primary cells. I used hTERT alone without success. Now I am usingLenti SV 40Tt and then Lenti HPV 16 E6/E7. With each of these treatments I am coupling them with Lenti hTERT. I was wondering if people had recommendations on methods for using hTERT with Lenti SV 40Tt or HPV 16 E6/E7? What are some immortalization tips that you might have?
For most cell types, hTERT expression is sufficient to immortalized cells. Since you did not mention whether hTERT was expressed detectably or not in your cells in your attempt to immortalize cells, it is harder to suggest whether you would need both SV40 large T and hTERT. One needs to consider potential problems in immortalizing cells with viral oncoproteins because a number of studies involving the pRb and p53 pathways can not be performed.
hTERT is not expressed detectably in the cells before immortalization.
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