Using genetic and molecular techniques to study anthocyanin production and protein expression in recombinant Escherichia coli
MetadataShow full item record
This dissertation involves the construction of the anthocyanin biosynthetic pathway into prokaryotic Escherichia Coli by combining genetic and molecular techniques. E.Coli has been frequently used as an expression system for the production of diverse classes of high-value compounds owing to its well studied genetics and growth characteristics. Anthocyanins belong to the wide class of flavonoids which are naturally produced as secondary metabolites in plant and have been a group of compounds which have generated incredible interest in the recent past owing to various health benefits including anti-oxidant and anti-diabetic effects. The first part of this thesis deals with the idea of improving protein expression and solubility to study its correlation with production of anthocyanins. Different plant sources such as Fragaria Ananassa and Vitis Vinifera were used for gene sequences encoding the enzyme 3-O-Glucosyltransferase (3GT) which was identified as limiting the reaction. Later strain engineering of E.Coli has been done in order to improve production levels with the idea of overexpressing 3GT so as to increase its gene dosage in the cell and increasing intracellular concentrations of UDP Glucose. Optimization of induction of protein expression was also carried out by inducing the cells during the later part of the log phase of their growth. Both these methods result in an increase in production and this opens up the scope of high yield production of anthocyanin in engineered E.Coli.