How to backtrack to DNA sequence from protein - (Aug/08/2007 )
I have a transgenic plant with a new transcription factor inserted. The transcription factor has turned-on some other protein (~45kDa) that isn't in the control plants. It would be really interesting to find out what the protein is. When I run an SDS-PAGE gel I can clearly see the band - so its in fairly high concentration.
What's the best way to backtrack from having the protein to getting the DNA sequence so I can blast it against some relative databases?
why do you need the DNA sequence for that? just use 'blastp' for protein sequences
I don't know what the protein is. All I know is that it is now turned on. I was wondering if cutting it out and sequencing it seems to be the right next move?
I would think I wouldn't have to sequence the entire protein, but enough to do a blast.
Yeah, cut it out and sequence it.
Or try mass spectrometry. Problem is if it isn't pure you may have alot of other stuff in with it but if it's that abundant hopefully it won't matter.
Since your transgenic plant is over-expressing a transcription factor you could use some kind of CHIP approach to IP the transcription factor bound to promoter sequences and then sequence the DNA sequences and find out potential genes it could be. This approach has been used for STAT1 target genes recently in one of the Naure journals or Science I think.