Hi Anamika, I did a search and it looks like researchers are using either mammalian cells or baculovirus to express this protein. I don't have experience with this particular construct so could you tell me why you choose *E. coli*? I run a protein expression facility and we typically use HEK or CHO cells and get mg amounts from cell culture of mammalian derived proteins like STAT1. Happy to talk offline if that would be easier.
Best, David -- David L. Blum, Ph.D. Director, Bioexpression and Fermentation Facility Department of Biochemistry and Molecular Biology University of Georgia 120 Green Street room A414A Athens, GA 30602 http://bff.uga.edu/ (706) 542-1035 (Office) On Thu, Jul 6, 2017 at 10:11 AM, Anamika Singh <anamika.ii...@gmail.com> wrote: > Hi, > > Is anyone has worked with STAT1 proteins? > > I have cloned the SH2 domain of STAT1 protein into pet28a vector but there > was no expression so far or rather say inconsistent expression. Sometimes > the expression was in inclusion bodies. I have tried different methods to > pull out the protein from inclusion bodies using urea, guanidium chloride > tween20 but none of them worked well. The yield was very low (very faint > band on SDS-PAGE ) from 3-liter culture. I changed the host cells from BL21 > to Rosetta DE3 cells but no success so far. > > We thought to use some other vector system like with SUMO tag but did not > proceed because the aim of the project to design inhibitor and tag will > interfere. > > Please suggest me something so that I can complete my project in lesser > time. > > > Looking forward to valuable suggestions. > > Thanks > Anamika >
