ISSN:
1617-4623
Keywords:
Yeast
;
HSV TK
;
Promoter
;
Interferon
;
Secretion
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Summary A system is described that facilitates the selection of yeast promoters of unknown origin using a functional heterologous gene assay. A multicopy plasmid was constructed with a unique cloning site 5′ to the Herpes Simplex type 1 thymidine kinase gene (TK). A yeast genomic DNA library was introduced into this site and used to transform yeast (which naturally lacks a TK gene). Only transformants carrying DNA fragments with promoter activity can express TK activity and thereby grow on medium containing thymidine and folate antagonists. On the basis of TK assays of yeast cell homogenates from some 100 TK + transformants, a particularly promising plasmid containing a yeast promoter was selected for further analysis. The position of the transcriptional start of the mRNA of this promoter was identified and the promoter sequence established. A fragment containing the promoter was taken and engineered to produce an expression vector and an expression/secretion vector. We report on the expression and secretion of interferon —X430 (IFN-X 430) (a derivative of IFN beta) from yeast cells containing such plasmids.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00331032
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