utilizing a transposon incorporated with luciferase for cancer studies, how do i distinguish between cells that form tumor and those that do not. As the transposon would have passed the luciferase gene to any cell it hit. I guess all cells hit by the transposon would have the same luciferase expression level. But how do i distinguish tumor from non-tumor cells.
Thanks
You can do immunostaining for cancer specific marker and colocalise it with luciferase expression.
All the best,
Jagadeesh
I would be careful about assuming the same level of luciferase activity. Depending on where the transposon integrates and how it is silenced by the genome, you could see different levels of expression. For example, see Garrison 2007 (http://mcb.asm.org/content/27/24/8824.full) for silencing effects in the mammalian genome.
For a first pass screen, you can do soft agar invasion assays, Ki67 staining etc to check for tumor forming properties of the transformed cells. But the best way is likely to inject the cells with the mutations into mice and see if they form tumors. David Largaespada's group has develop a lot of a great tools for forward screens of cancer genes using transposon systems.
@Jagadeesh thanks lot. i actually can stain with cancer markers, but i dont understand what you mean by colocalizing with luciferase activity.
@Fiorella Grandi. Thanks very much for your point. I can do Ki67 staining to check which of the formed cells are transformed cells.
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