I am planning a new set of experiments and would like to use the gateway cloning system. At this point I only want to test a few sequences for regulatory potential (but could see this going to a much more high-throughput screen). I want to make this process efficient with as few steps as possible. After reading up on the system, I am left wondering if you could engineer a final "destination" plasmid (your reporter construct) that contained the attp1 and attp2 sites flanking a positive and negative selectable marker. Then simply clone PCR fragments flanked by the attB1 and attB2 sequences. Essentially this would be the same as creating the entry vector, I believe. Does this seem like it would work. It almost seems too simple and I'm surprised that I haven't already seen this protocol somewhere and makes me think I am missing something. Thoughts? Thanks!!!
Edited by lmh327, 31 March 2014 - 06:28 AM.