Executive Summary : | Polyunsaturated fatty acids (PUFAs) are essential for human health, but humans cannot produce them due to their inability to obtain them from the diet. To meet this demand, extensive research is being conducted to produce LCPUFAs from alternative sources, such as deep-sea fish oil, peanuts, soybeans, and flax seeds. However, extracting LCPUFAs from plants is laborious and unsustainable. Unconventional yeasts, such as Kluyveromyces marxianus, have been used as an alternative to Escherichia coli and Saccharomyces cerevisiae due to their broad range of sugar utilization, rapid growth rate, and thermotolerant nature. This research aims to isolate and identify key genes involved in the PUFA biosynthetic pathway from microalgae Isochrysis sp., and engineer them in K. marxianus to produce LCPUFAs.
The study will use synthetic biology tools to confirm gene insertion, analyze protein expression, demonstrate functional characteristics, and estimate total lipid content using Nile Red Fluorescence assay. Cultural conditions, including substrates, temperature, and time, will be applied to maximize the production of fatty acids by the recombinant K. marxianus. Previously, fatty acid production was reported in E. coli and S. cerevisae, but these studies focused on producing PUFAs by expressing one or two genes. This study aims to produce economically valuable VLCPUFAs by engineering five fatty acid biosynthetic genes in K. marxianus, bringing a breakthrough in fatty acid synthesis and revolutionizing human society to prevent and cure neurodegenerative diseases. |