The i-motif is a DNA quadruplex structure of interest for its potential to modulate gene expression. This project will optimise the conditions required to crystallise an intramolecular i-motif.
Department: Pharmacy
Supervised by: John Brazier
The i-motif is a four stranded DNA structure that can form in DNA sequences of biological interest. Recent studies have shown that quadruplex structures in the promoter region of genes can play a role in gene expression, as such these structures provide an attractive therapeutic target. Currently information regarding the structure of these quadruplexes is limited to crystallographic intermolecular structures of small DNA sequences or NMR solution structures of longer intramolecular sequences. This project aims to optimise the conditions required to crystallise longer intramolecular sequences to be studied by X-Ray diffraction. The crystals prepared in this project will be used to generate a detailed structure of this interesting quadruplex structure. Detailed structural information will guide the design of small molecule ligands capable of stabilising formation of the i-motif leading to therapeutic applications. To date no intramolecular i-motif crystal structures have been reported, further emphasising the need for this research.
The student will perform hands on experiments in a laboratory setting. This will include handling of oligonucleotides, preparation of crystallisation solutions and experiments. The student will work under the guidance of James Hall and Prof Cardin, preparing crystallisation trays using a range of conditions. These 24-well trays are set aside for possible crystallisation, a process which may take days or weeks. The trays are observed at intervals of a few days under the microscope for signs of crystal growth. If even small crystals are obtained, they can be studied on the microfocus beamline I04 at Diamond, as James Hall is a Diamond-funded student, and and has regular access to the macromolecular beamlines.
Experience of conducting experiments within a laboratory would be desirable/ essential. Background knowledge of DNA structure and determination of structure by x-ray diffraction would be advantageous. For setting up the crystallisation trials, the skills required are those for working on the microlitre scale – pipetting, clean working techniques, patience, and careful observation. The oligonucleotides used are expensive, so the project is suitable for a careful, methodical and systematic approach. Some basic physical chemistry, up to first year undergraduate level, is very helpful.
The student will gain several laboratory skills, including handling of oligonucleotide samples and use of micropipettes. Preparation of crystal samples and subsequent analysis of suitability for x-ray diffraction studies will help to develop both general research experience and analytical skills. Working with two supervisor will develop the students communication skills, especially with regard to scientific discussion. Successful growth of crystals for x-ray diffraction analysis would result in authorship of subsequent structure papers.
Chemistry Building
9-5
Monday 09 July 2012 - Monday 20 August 2012
Students should submit a CV and covering letter when applying for this project. Interviews will be held to assess the suitability of the students but not formal testing will take place.