The development of Xanthophyllomyces dendrorhous as a renewable source of high value carotenoids. / Ali, Thunazzina.

2020. 332 p.

Research output: ThesisDoctoral Thesis

Unpublished

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  • The development of Xanthophyllomyces dendrorhous as a renewable source of carotenoids

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Abstract

In the food, pharmaceutical and aquaculture industries, astaxanthin, a reddish carotenoid is
typically the most expensive component. A sustainable, economically viable and non-GM
source of carotenoids has been developed in Xanthophyllomyces dendrorhous, a
basidiomycetous yeast that produces astaxanthin. The fully sequenced genome of X.
dendrorhous has been manipulated through chemical mutagenesis for untargeted strain
improvement using N-methylnitronitrosoguanidine.
Second-generation mutants have produced competitive levels of various carotenoids, when
compared to their first-generation counterparts. Mutant strain Eg60/Eg34 produced a 7-fold
increase in astaxanthin compared to the wild-type. Other mutant strains such as Eg27/Eg15
produced a 100-fold increase (370 µg/g DW) of phytoene and Eg24/Eg15, an 11-fold increase
(388 µg/g DW) of β-carotene. Gene expression experiments were undertaken by using qPCR
and indicated that the elevated carotenoid levels regulate the carotenoid pathway via a
negative feedback loop. Single nucleotide polymorphisms (SNPs) have been identified in
candidate genes of non-carotenoid, phytoene and β-carotene producers, in phytoene
synthase (CrtYB), phytoene desaturase (CrtI) and astaxanthin synthase (asy) respectively.
These detrimental SNPs have created blocks within the pathway and have caused changes in
the overall metabolism between strains.
Metabolite profiling conducted via gas chromatography - mass spectrometry (GC-MS)
illustrated differences in carbon flux between first and second-generation mutants. Both
phytoene and β-carotene second round mutants had increases in other metabolite families
such as the amino acids and fatty acids respectively. Lastly, experiments conducted to
understand the cellular and physiological changes within mutant cell lines, demonstrated
large differences in structures against the WT. Therefore, proposing new structures and
metabolites were necessary to accommodate for the alterations in carotenoid content.
Clean mutants can be used as a background to further investigate strains produced by
chemical mutagenesis. Therefore, the genotypes created can be used as a chassis or in
combination with CRISPR-Cas as a method of targeted pathway engineering for strain
improvement.
Original languageEnglish
QualificationPh.D.
Awarding Institution
Supervisors/Advisors
Award date1 Jul 2020
Publication statusUnpublished - 13 Jun 2020

ID: 38288447