DNA in the cell is tangled and twisted, adopts complex topologies and is frequently maintained under superhelical stress. There is now a growing appreciation that the context of DNA in the wider genome is vital to its function and can affect many interactions key to cell viability. Understanding how DNA behaves in its cellular environment is currently a challenge of complexity, limited in part by the biophysical tools available. Our lab uses single-molecule Atomic Force Microscopy (AFM) methods to observe local variations in the structure, conformation and dynamics of individual DNA molecules and associated binding proteins at sub-nanometre spatial resolution and sub second temporal resolution [1]–[3].
DNA structure at double helical resolution - Pyne et al., Small (2014)
Your interdisciplinary project will use a combination of cutting edge biophysical techniques and gold-standard biochemical assays to determine the effect of complex DNA topologies and supercoiling on DNA structure, with double-helical resolution. Your project will look to explicitly determine how DNA responds to supercoiling and stress, and how this influences the interaction of DNA with regulatory proteins, sensitive to these variations in structure. Your project will build upon our expertise in high-resolution Atomic Force Microscopy, to visualise variations in DNA structure and conformation, only observable when molecules are probed on these time and length scales. You will enhance this data through additional cutting-edge single-molecule techniques at the University of Sheffield [4] (www.craggs-lab.com). To evaluate these interactions, you will work with us to develop our quantitative, open-source analysis tools to exploit our large datasets, providing unique insights into biomolecular structure and function that are essential to life.
Environment:
You will be part of an interdisciplinary team, working to address biological questions using physical techniques. You will be primarily supervised by Dr Alice Pyne (Dept. of Materials Science) with expertise in high resolution AFM of biomolecules (www.pyne-lab.uk). We collaborate internationally with academia and industry across the sciences and engineering, with applications from fundamental sciences to novel therapeutics. You will join a collaborative, supportive research community at Sheffield, with world-leading single molecule and nucleic acid research centres (http://www.imagine-imaginglife.com and genome.sheffield.ac.uk respectively) and an active, friendly and lively PhD student cohort. We are committed to supporting the career development of our students, encouraging attendance at both international and UK meetings, conferences and training courses to develop your research skills and interests.
About you:
As an interdisciplinary project, we welcome applicants from a diverse range of backgrounds across engineering and the physical and biological sciences. Interested applicants should contact Alice to discuss the project further (a.l.pyne@sheffield.ac.uk).
Start date: 28th September 2020 (negotiable)
Good luck!
Funding Notes:
This is an EPSRC studentship, funded for 3.5 years including academic fees at the UK/EU rate (currently £4,329 per year for 2019/20) and a stipend at EPSRC rates (£15,009 p.a.). To qualify, you must be a UK or EU citizen who has been resident in the UK/EU for 3 years prior to commencement. Applicants must have obtained, or be about to obtain, at least a 2.1 honours degree (or equivalent) in a relevant subject.
References:
[1] A. Pyne, R. Thompson, C. Leung, D. Roy, and B. W. Hoogenboom, ‘Single-Molecule Reconstruction of Oligonucleotide Secondary Structure by Atomic Force Microscopy’, Small, vol. 10, no. 16, pp. 3257–3261, Apr. 2014.
[2] B. Klejevskaja et al., ‘Studies of G-quadruplexes formed within self-assembled DNA mini-circles’, Chem. Commun., vol. 52, no. 84, pp. 12454–12457, Jan. 2016.
[3] B. Akpinar, P. Haynes, N. Bell, K. Brunner, A. Pyne, and B. Hoogenboom, ‘PEGylated surfaces for the study of DNA-protein interactions by atomic force microscopy’, Nanoscale, p. 10.1039.C9NR07104K, 2019.
[4] B. Hellenkamp et al., ‘Precision and accuracy of single-molecule FRET measurements—a multi-laboratory benchmark study’, Nat. Methods, vol. 15, no. 9, pp. 669–676, Sep. 2018.