Eastern Kentucky University
Lipid Production in Chlorella Protothecoides
Institution
Eastern Kentucky University
Faculty Advisor/ Mentor
Rebekah Waikel
Abstract
Given the concern for diminishing fossil fuel supplies and the environmental consequences of fossil fuels, it is vital to seek out alternative energy resources. Our laboratory seeks to study the use of the algal species, Chlorella Protothecoides, in biofuel production by identifying the genes that control lipid production as well as their regulation. To identify genes that are differentially regulated in lipid producing environments, we performed Next Generation Sequencing and Real Time PCR on algae grown in growth conditions (nitrogen sufficient) compared to algae grown in lipid producing conditions (nitrogen deficient). Lipid production in nitrogen deficient conditions was confirmed using Bodipy 505/515 staining, analyzed by flow cytometry. Analysis of Next Generation Sequencing data yielded distinct algal gene expression profiles for both lipid producing conditions and growth conditions. Blast analysis of differentially regulated mRNA sequences revealed the identity to known genes, as well as several unknown genes. Known genes thought to be involved in lipid production and eight (8) yet to be identified genes were further analyzed by quantitative Real Time PCR. We have confirmed that dihydrolipamide acetyltransferase and acetyl transferase are upregulated in lipid producing conditions. We have also confirmed the differential regulation of the 8 unknown genes. These confirmed differentially expressed genes will now be used as lipid producing markers to identify environmental condition such as pH, light, micronutrients, temperature, etc., which optimize lipid production.
Lipid Production in Chlorella Protothecoides
Given the concern for diminishing fossil fuel supplies and the environmental consequences of fossil fuels, it is vital to seek out alternative energy resources. Our laboratory seeks to study the use of the algal species, Chlorella Protothecoides, in biofuel production by identifying the genes that control lipid production as well as their regulation. To identify genes that are differentially regulated in lipid producing environments, we performed Next Generation Sequencing and Real Time PCR on algae grown in growth conditions (nitrogen sufficient) compared to algae grown in lipid producing conditions (nitrogen deficient). Lipid production in nitrogen deficient conditions was confirmed using Bodipy 505/515 staining, analyzed by flow cytometry. Analysis of Next Generation Sequencing data yielded distinct algal gene expression profiles for both lipid producing conditions and growth conditions. Blast analysis of differentially regulated mRNA sequences revealed the identity to known genes, as well as several unknown genes. Known genes thought to be involved in lipid production and eight (8) yet to be identified genes were further analyzed by quantitative Real Time PCR. We have confirmed that dihydrolipamide acetyltransferase and acetyl transferase are upregulated in lipid producing conditions. We have also confirmed the differential regulation of the 8 unknown genes. These confirmed differentially expressed genes will now be used as lipid producing markers to identify environmental condition such as pH, light, micronutrients, temperature, etc., which optimize lipid production.