I am doing cloning with the pETDuet1 construct. The original pETDuet1 and the pETDuet-1 with my insert ( approx 1 kb) are transformed in DH5alpha cells. I have noticed that my mini preps have very low yield (= 50 ng/ul) and I get no pellets in my midi preps! i have alternately tried the Qiagen and Macherey Nagel kits and I know the kits are fine. My midi preps are using a colony from a freshly streaked plate and inoculated into 200 ml LB-AMP, grown O/N (= or < 16 hrs) at 37C. My ampicillin stocks are fresh and I use Amp at a final conc of 100ug/ul. Has anyone had a similar problem with pETDuet1 plasmid?
Plasmid preps of pETDuet-1 constructs
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