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cloning of cellulose synthase gene - (Jan/24/2010 )

Dear friends
Im trying to clone a gene encoding a cellulose synthase from agrobacterium into the expression vector pET 30b. The molecular weight of the protein encoded by the gene is 150 kDa as calculated from the protein sequence. Is it possible to express this big protein in e.coli?and are there any assays for cellulose synthase?pls help

-chn09-

chn09 on Jan 24 2010, 05:28 PM said:

Dear friends
Im trying to clone a gene encoding a cellulose synthase from agrobacterium into the expression vector pET 30b. The molecular weight of the protein encoded by the gene is 150 kDa as calculated from the protein sequence. Is it possible to express this big protein in e.coli?and are there any assays for cellulose synthase?pls help


Hi, it should not be a big issue expressing protein of this size in E.coli....ppl do express proteins of about 400 kDa........may degrade much more rapidly on repeated freeze and thawing..........also just take care of codon bias if any.....for the assays..........these references might help....

1. Blanton R L(1993) Development (Cambridge, UK) 119:703710.


2. Wong H C, Fear A L, Calhoon R D, Eichinger G H, Mayer R, Amikam D, Benziman M, Gelfand D H, Meade J H, Emerick A W, et al.(1990) Proc Natl Acad Sci USA 87:81308134, pmid:2146681.


Best...

-DRN-