I'm trying to anticipate how many mice I'll need for RT-qPCR of several genes expressed in the midbrain, specifically the VTA. I need to be able to get about 4mg of VTA tissue from each brain. Is this a reasonable expectation?
Hi Clayton. We use a single midbrain (from adult mice) for our RTqPCR analysis, though we generally include the entire midbrain. A rough estimate would be that we end up with 10mg. But the amount of tissue you need depends on the specific genes you need to test. Good luck:)
We routinely micro-dissect areas of the brain 1-2mm thick, and are weighed in micro grams. 4 milligrams is a lot of tissue, I doubt that you will achieve 4mg that from just the VTA in a single mouse.
Hi Clayton,
I agree with Jeffrey, 4 mg of VTA from one adult mouse is a lot.
On top of that, you need to be rather skilled to dissect out the VTA as I find it not an easy structure to discern/dissect. Some take the whole midbrain, like Freja indicated.
If you go for the VTA alone; you may want to consider a conservative excision, thereby ending up with 2-3 mg from which you are sure you have VTA-tissue and pool VTA from two mice? TH gene expression could be employed to ensure the selectivitiy of your excision method?
Good luck,
Michael