2 replies to this topic

[scary]

Hi

I am trying to express my glutamate synthase gene in bacterial system. But, I do not have sequence of complete gene so I was wondering if i can express the protein from partial sequence??  If it is possible to express the protein, which vector system would be appropriate? my ultimate goal is to raise antibodies against the purified protein.

I would really appreciate any suggestions. Thankx in advance.

-scary

[fishdoc]

scary, on Feb 21 2010, 11:04 AM, said:

Hi

I am trying to express my glutamate synthase gene in bacterial system. But, I do not have sequence of complete gene so I was wondering if i can express the protein from partial sequence??  If it is possible to express the protein, which vector system would be appropriate? my ultimate goal is to raise antibodies against the purified protein.

I would really appreciate any suggestions. Thankx in advance.

-scary

You can express it, but there's no guarantee a truncated protein will be active. For antibody production, activity probably isn't necessary. For the vector, any high copy number vector should be sufficient. Most of them will have some sort of inducible promoter to express the protein at high levels in E. coli. We use pBluescript in our lab, but I think it's kind of an outdated vector.  There are a number of commercially available TOPO kits available for such expression. Just need to find one that suits your needs and your budget. In most cases, you can fuse the open reading frame to a epitope tag (6x His) that will be useful for getting the protein purified for antibody production.

[scary]

hi fishdoc

thanks for clearing my doubt.