Abstract

The ATP gated cation channel P2X7, is a cell surface protein whose function and activation is linked with several human diseases. The aim of this project was to raise DNA Aptamers targeting specifically the extracellular domain of the P2X7 protein with the hope to use these new ligands as a method for the study of the biological functions of the P2X7 protein and its potential as a therapeutic target. The strategy involved isolating Aptamers that would bind to human embryonic kidney cells transformed with a construct which forced the cells to express native mouse P2X7 on its cell surface (POSITIVE SELECTION), whilst not binding to untransformed non-P2X7 expressing cells (NEGATIVE SELECTION). Two candidate sequences from this selection emerged as, what we called, "winners" which were the most likely sequences to putatively bind mP2X7 (work ongoing). Interestingly using the procedure in reverse, that is to create Aptamers that specifically bind to non-transformed cells and not transformed cells, also produced viable putative Aptamers, which, once labelled, showed selective binding. These results are discussed in relation to future applications and the potential insights and implications for the study of cell surface protein expression patterns.

Additional Notes

Supervisors: Dr James McClellan and Dr Darren Gowers

Course: Master of Philosophy - MPhil

Date Deposited: 2017-05-12

URI/permalink: https://library.port.ac.uk/dissert/dis12887.html