[Chlamydomonas] colony lifts

Maureen Wirschell via chlamy%40net.bio.net (by wirsch from cellbio.emory.edu)
Tue Feb 27 14:03:04 EST 2007


Has anyone has tried doing colony lifts on Chlamy?  (Like how we do colony lifts for bacterial colonies or phage lifts for phage screening?).  I was looking for a quick higher throughput method for screening for RNAi transformants, other than to try westerns on whole cell lysates or axonemes.  We have about 1000 transformants (co-transformed with either pHyg3 for hygromycin resistance, or pARG7.8 for arginine) that we need to screen for RNAi of our gene.  

Our attempts at colony PCR are inconsistent--even getting consistent PCR with mating type primers seems to be a bit troublesome in our hands, so we typically use purified genomic DNA for consistent PCR.  So, I got to thinking about doing some type of colony lift and I don't know if this has been tried with Chlamy at all.  Just wondering.  

I may opt to redesign my RNAi construct to put in the pHyg cassette for hygromycin resistance, so I can do direct selection.  Nonetheless, I still got to thinking about whether or not anyone has even tried doing southerns on colony lifts from Chlamy plates.  

If anyone has any thoughts, that would be great.
Maureen Wirschell




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