Epsilon Archive for Student Projects

Abstract

When starch is produced, it is often subjected to chemical treatments in order to achieve desired characteristics. Today, it is possible to genetically modify the potato genome using the CRISPR/Cas9 technique to influence starch synthesis and produce potatoes with a desired starch composition and structure. This enables the production of starch without the need for chemical modification by genetically modifying enzymes involved in starch biosynthesis, such as starch synthases (SS), starch branching enzymes (SBE), and starch debranching enzymes (DBE). To further study these enzymes, one approach is to express them in Escherichia Coli. This system allows for a cost-effective production of high protein yields and opens the door for enzyme engineering to tailor or enhance specific catalytic properties. These insights can later be utilised for more accurate editing of the enzymes in potato. This study has utilized a gateway cloning system from Invitrogen, in which SS1 has been transformed into E. coli via the donor vector pDON221 and the destination vector pDEST17. Followed by attempts to induce protein expression with IPTG. This study confirmed the insertion of the SS1 in E. Coli, but no confirmation of expression was achieved. To express functional proteins in E. coli, several optimization steps during the induction of expression need to be taken into consideration. This study provides a solid base for further attempts to express and obtain soluble active SS protein in E. Coli.