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Lookup NU author(s): Mark Pearson, Dr Siavash Khazaipoul, Professor John Hesketh
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Chinese hamster ovary (CHO) cells are used for recombinant protein production in the pharmaceutical industry but there is a need to improve expression levels. In the present work experiments were carried out to test the effectiveness of different 3'untranslated regions (3'UTRs) in promoting production of a naturally secreted luciferase. Seamless cloning was used to produce expression vectors in which Gaussia princeps luciferase coding sequences were linked to the human albumin, immunoglobulin or chymotrypsinogen 3'UTR. Stably transfected CHO cells expressing these constructs were selected. Luciferase activity in the culture medium was increased 23-fold by replacing the endogenous 3'UTR with the albumin 3'UTR and decreased by replacement with immunoglobulin or chymotrypsinogen 3'UTR. Replacement of the native 3'UTR with the albumin 3'UTR led to a 10-fold increase in luciferase mRNA levels. Deletion analysis of the albumin 3'UTR showed that loss of nucleotides 1-50, which removed an AU-rich complex stem loop region, caused significant reductions in both luciferase protein expression and luciferase mRNA levels. The results suggest that recombinant protein expression and yield could be improved by the careful selection of appropriate 3'UTR sequences.
Author(s): Pearson MJ, Khazaipoul S, Optun A, Pryme IF, Stern B, Hesketh JE
Publication type: Article
Publication status: Published
Journal: Biotechnology Journal
Year: 2012
Volume: 7
Issue: 11
Pages: 1405-1411
Print publication date: 29/06/2012
ISSN (print): 1860-6768
ISSN (electronic): 1860-7314
Publisher: Wiley - V C H Verlag GmbH & Co. KGaA
URL: http://dx.doi.org/10.1002/biot.201200044
DOI: 10.1002/biot.201200044
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