Construction of an expression vector for the fission yeast Schizosaccharomyces pombe.

Abstract:

:We have isolated and characterized a S. pombe promoter using a functional heterologous gene product assay. Random S. pombe genomic fragments were cloned upstream from the promoterless 'lacZ gene and tested in vivo for their efficiency to promote expression of the beta-galactosidase protein in the fission yeast. An efficient S. pombe promoter called 54/1 was isolated and shown to drive up to 5% of total protein synthesis as beta-galactosidase. The structure and nucleotide sequence of this promoter were determined, precise localization of its mRNA transcriptional start points established. Translational fusion of the Pseudomonas putida XylE gene with the 54/1 gene was shown to allow expression of catechol oxidase activity in S. pombe. An expression vector suitable for transcriptional fusions was then constructed from engineered 54/1 promoter sequences and used to drive expression of the E. coli Tn5 ble gene, thus confering resistance to the fission yeast against bleomycin and phleomycin antibiotics.

journal_name

Nucleic Acids Res

journal_title

Nucleic acids research

authors

Kudla B,Persuy MA,Gaillardin C,Heslot H

doi

10.1093/nar/16.17.8603

subject

Has Abstract

pub_date

1988-09-12 00:00:00

pages

8603-17

issue

17

eissn

0305-1048

issn

1362-4962

journal_volume

16

pub_type

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