ISOLATION AND CLONING OF PUTATIVE PROTEIN SACYLTRANSFERASE (PAT) GENE OF MARINE MICROALGAE CHLORELLA VULGARIS

• Main Author: AYU PRANA (NIM:10514011), RISNA
• Format: Final Project
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/30539
• Institution: Institut Teknologi Bandung

World’s fuel consumption is increasing year by year, but it is not comparable to the amount of <br /> <br /> production. An alternative renewable fuel beside fossil fuel is needed. Microalgae has great potential as a <br /> <br /> renewable fuel source because microalgae are the highest accumulated lipid-producing microorganisms, <br /> <br /> which is about 20-50% of the weight of dry biomass. To increase the economic value of fuel from <br /> <br /> microalgae, we need to increase the lipid production from microalgae by genetic engineering of lipid <br /> <br /> biosynthesis pathways in microalgae. S-acyltransferase protein (PAT) is one of the proteins that has an <br /> <br /> important role in lipid biosynthesis. This protein is a key in the process of transfer of acyl chains to <br /> <br /> acceptor molecules in TAG biosynthesis. Without these proteins, the cycle of lipid biosynthesis will stop <br /> <br /> until the formation of fatty acids. This research aims is to isolate and clone the S-acyltransferase gene <br /> <br /> from Indonesian marine microalgae, Chlorella vulgaris. The research stages included the preparation of <br /> <br /> pure microalgae culture, microalgae activation and cultivation, investigation of cell morphology under the <br /> <br /> light microscopy, total DNA isolation, amplification of PAT genes fragment with PCR, sequencing of <br /> <br /> PAT gene fragment, total RNA isolation, RT-PCR for synthesis of the first cDNA chain, PCR on the first <br /> <br /> cDNA chain using spesific primers of PAT gene, cloning PAT gene fragments using pGEM-T vector and <br /> <br /> E coli TOP 10F’ as host cell. This study research, 0,7 kb and 0,9 kb fragments of PAT gene have been <br /> <br /> successfully cloned in E. coli cells.