Tet resistant colonies will not grow in medium - (Mar/16/2005 )
I am trying to make some BL21 AI (arabinose inducable) competent cells to use in protein purification. ( I have a small stock of cells frm a lab we are collaborating with.) The competent cells, according to Invitrogen, have a tetA resistance gene so I can use tet to confirm strain identity. When I streak a small amt of cell stock onto LB/tet 10 ug/mL plates, I get single colonies, but when I seed a single colony from this plate into 2mL LB/Tet 10 media and shake O/N 250 prm 37C, I get no growth! So it seems the colony I picked, even though it was from the Tet plate, is not tet resistant! What am I missing/overlooking here? Thanks.
This might sound weird but inoculate liquid media with Tet directly from the stock instead of picking up one colony. Carry on with the rest of your expts. Sounds strange? It will work.