cloning problem

[Bryan J. Maloney]

Britta Satzke <bsatzke at gwdg.de> wrote in news:3D91CA72.B2287E2 at gwdg.de:

> Hi, there
> 
> all lab members have had some cloning problems for two weeks now:
> 
> 1. When subcloning in DH5a we get 2 types of colonies: big and small
> ones.
> 
> 2. After Mini-DNA-prep we always get two bands (size 3kb + 1.8kb)- no
> matter which vector and insert we use, nor which restriction enzymes we
> use.
> 
> We get low transformation efficiencies with Midi-DNA, transformation of
> ligation doesn´t work anymore.

You have a bacterial contaminant that has overwhelmed your culture.  Go 
back to clean DH5a stock.