I'd like to use this assay to detect fragmentation of a large viral genome, but don't know if those fragment sizes can be distinguished using this approach.
The comet assay cannot provide any information about DNA fragment size, because the electrophoresis step is too short to allow for any real separation. Running the electrophoresis for too long will cause false positive tails on the control cells. It's better to perform a pulse field if you need to know specific fragment lengths. Here's an article that explains this and more about the comet assay: http://atm.skkumed.ac.kr/protocol/comet%20assay%202008%20nature%20protocols.pdf
Thanks. I don't have direct experience with this assay, so I appreciate your insights. I agree that direct measurement of fragment size isn't possible, I was hoping this method would be sufficient to separate 1 fragment from another 4 times as long in the size ranges I gave... it doesn't sounds like that is feasible though. PGE would be ideal, but the necessary apparatus isn't cheap, I was hoping that a comet assay could provide a cruder estimate of whether DNA damage was occurring to justify buying a PGE unit.
Again, thanks for your time and for the protocol.
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