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[anonymous]

hello everybody!!                          I am  unable to clone HIV pol gene. My insert get deleted on cloning. It                          becomes 300bp from 3kb,                           I am thankful to those who suggested me to try different host strains but it                          has not helped so far. Just to remind , I am using Topo Cloning kit and I have                          tried Topo 10 cells, Life tech max Stbl 2 cells Stratagene SURE 2 cells anf                          HB101 cells, but still I have the same problem.                          many thanks on your much anticipated suggestions.

[anonymous]

I had a similar problem.  I ended up having to use Invitrogen's pBAD topo cloning system to shut down expression.  Adding glucose to the overnight culture was essential too.