Overexpression and Purification of the Oat (Avena Fatua) Protein AFN1 and Construction of a pMAL/AFN1 Expression Plasmid

Abscisic acid (ABA) is an important phytohormone with regulatory roles in many physiological processes. ABA expression is induced by environmental stresses such as drought and it is known to be an inhibitor of seed germination. A wild oat (Avena fatua) called AFN1 has been hypothesized to initiate the early stages of germination as its mRNA accumulates in nondormant seed embryos during imbibition. The polypeptide sequence of AFN1 suggests that it is an ABA glucosyl transferase. Glucosylation by AFN1 and thereby inactivation of ABA could lead to seed germination. In order to understand the role of AFN1 in germination, an ample quantity of AFN1 polypeptide is needed to test for enzymatic ABA glucosylase activity. My work has been to overexpress recombinant AFN1containing a (His)6 tag using a pRSETC E.coli expression system followed by Purification of the AFN1 protein by means of a nickel-affinity column that bind to the (His)6 tag. Due to the insufficient yield of AFN1 fusion protein obtained with this procedure, another method using a pMAL-c2x vector is now being employed. The pMAL expression system provides a method for expressing and purifying protein by tagging proteins with maltose-binding protein (MBP). It is anticipated that MBP tag will be advantageous as it can make the fusion protein more soluble and thereby yield a larger quantity of protein. Currently, work is underway on the construction of pMAL/AFN1 plasmid.

'Off with his head!' : Inexplicable Faith in Bulgakov's Master and Margarita

Examining how, in the novel, Bulgakov shows the conflict between logic and faith through the actions of his characters : the characters who are logical are generally not portrayed as wise and are said to not appreciate, nor understand, faith.

Art of Goya as a significant reflection of his time

This thesis examines the story of how Goya came to be the artist he was and of the times which made him what he was, and which he so well mirrored. Francisco Goya y Lucientes is a remarkable figure in the history of art. Rarely has an artist arisen from such a cultural vacuum; rarely has an artist done so much to establish a cultural reputation for his country; rarely has an artist reflected so accurately the spirit of his time; and rare is the artist who has such an important influence on all the significant art of the future.