Abstract
A variant of Saccharomyces cerevisiae pep4 strain 20B12, with improved oligotrophic proliferation, cell survival and secretion of heterologous mouse α-amylase, is described. Previously we reported a procedure to enrich NI transformants that are not inhibited by cytotoxic expression of hepatitis B virus surface antigen in the secretion pathway of the protease-A-deficient (pep4) strain. To use the NI cells as a host for heterologous expression, we tried to amend the introduced pYAS/12S vector and obtain a host strain, NI-C, with stable NI phenotype and trp1 marker restored. Southern analysis of genomic DNA of NI-C suggested that the original pYAS/12S was abnormally rearranged and not completely corrected. Further assay showed that the viability and mitotic ability of the NI-C strain were increased. While using the NI-C strain as host for plasmid transformation and heterologous expression of mouse α-amylase, we observed that transformed colonies grew more quickly and secreted more α-amylase than general yeast strains. A further test showed that the NI-C strain was able to use mouse α-amylase as a positive selection marker to form transformed colonies on nitrogen-starved plates that contain starch as the sole carbon source. The results imply that the NI-C variant is an improved pep4 strain that can be used for heterologous expression and for the development of new selective markers in the yeast transformation system.
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Received: 7 January 1998 / Received last revision: 7 September 1998 / Accepted: 11 October 1998
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Chen, DC., Chen, SY., Gee, MF. et al. A variant of Saccharomyces cerevisiae pep4 strain with improved oligotrophic proliferation, cell survival and heterologous secretion of α-amylase. Appl Microbiol Biotechnol 51, 185–192 (1999). https://doi.org/10.1007/s002530051380
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DOI: https://doi.org/10.1007/s002530051380