Dear Prakriti

the only thing that you can do check if the problem is related to Genescript is sequencing your vector to confirm the the insert and the elements important for protein expression as promoter , rbs, stop codon, terminato are the expected.

just some questions?

is it a mammalian or eucariotick protein?

it was expressed soluble in the exact same form that you try? as n- or c term tags? domain?

ciao

Manuele

Hello Manuele Martinelli

Thanks for your reply. I am trying to get the plasmid sequenced and will know then.

It is a eukaryotic protein but it is only a small domain of the whole protein and should be around 18 kDa.

Yes, it has been expressed in the exact same form with N-terminal his-6 tag. The only difference is that my construct has a His-10 tag at the N-terminus.

Hi Prakriti, I order all my plasmids from Genscript and I have never had a problem with any of them expressing. Of course, there could always be a mistake, but I've always found their products to be great quality. Their customer service reps are also very responsive on email, so you could ask them for some more advice or information

Hi Prakriti,

I am also having serious problems with Genscript constructs. The genes are from a gram negative bacterium (in pET22 and pET30 vectors) and I am expressing them in E.coli BL21/C41 cells. I have always had great success with Genscript plasmids but something seems to be wrong with the 6 new constructs.

I have contacted the company and hoping to hear from them soon. My guess is that the codon optimization is problematic.