Investigation of the function and regulation of astaxanthin synthase in Xanthophyllomyces dendrorhous.

摘要

This thesis describes the study of the function and regulation of the crtS gene in X. dendrorhous and the expression of genes in the astaxanthin biosynthetic pathway.;Chapter 1 describes the yeast X. dendrorhous and its ability to produce astaxanthin. The function and importance of astaxanthin is then discussed followed by a description of the astaxanthin biosynthetic pathway in X. dendrorhous.;In Chapter 2, expression of the genes in the wild-type strain CBS6938 over time is studied by qRT-PCR. I also examined the synthesis of astaxanthin when the wild-type strain was grown under dark growth conditions and in the presence of light. Finally, I examined carotenogenic gene expression of the wild-type strain grown in dark and in light to determine if the differences in astaxanthin production are due to differences in gene expression.;Chapters 3 and 4 discuss the generation of a crtS knockout mutant strain and crtS repair strains to examine the role of the crtS gene in the conversion ofβ3-carotene to astaxanthin. I also characterized the function of crtS to determine if it can perform both ketolation and hydroxylation.;In Chapter 5, I examined gene expression in the knockout mutant strain and the two repair strains to further expand the knowledge of how the carotenogenic genes in X. dendrorhous are regulated. I compared gene expression in the knockout mutant strain and repair strains to the wild-type strain to determine if deletion of the crtS gene and insertion of the crtS cDNA affected gene expression.;In Chapter 6, I applied a procedure not used in the study of X. dendrorhous before to attempt to increase expression of the crtS gene to increase astaxanthin production. This involved incorporating a gapdh promoter from X. dendrorhous in front of the crtS cDNA and then reinserting into the crtS knockout mutant strain at the original crtS location.;Finally, Chapter 7 discusses the importance of the information gained through these studies and areas where further study is needed.