Alexandersson, Erik
- Lund University
Abstract
Conclusions: We report a convenient and robust method based on qPCR to determine recombinant gene dosage. The method is generic for all constructs based on the pPICZ vectors and offers an inexpensive, quick and reliable means of characterising recombinant P. pastoris clones. By using this method we show that: (1) heterologous expression of all aquaporins investigated respond strongly to an increase in recombinant gene dosage (2) expression from a single recombinant gene copy varies in an isoform dependent manner (3) the poor expression observed for AtSIP1;1 is mainly caused by posttranscriptional limitations. The protein folding and membrane localisation seems to be unaffected by increased expression levels. Thus a screen for elevated gene dosage can routinely be performed for identification of P. pastoris clones with high expression levels of aquaporins and other classes of membrane proteins.
Keywords
Pichia pastoris aquaporins; Major Intrinsic Proteins; qPCR
Published in
BMC Biotechnology
2011, volume: 11
Publisher: BIOMED CENTRAL LTD
SLU Authors
UKÄ Subject classification
Biochemistry
Molecular Biology
Publication identifier
- DOI: https://doi.org/10.1186/1472-6750-11-47
Permanent link to this page (URI)
https://res.slu.se/id/publ/40899