Improved gene editing by CRISPR/Cas9
This project will develop new cutting-edge technologies for the correction of inherited mutations responsible for all manner of human disease.
CRISPR-Cas9 is a powerful sequence specific DNA cutting enzyme that can be used to correct genetic mutations. However, Cas9 breaks are repaired by one of two major pathways in cells: Non-homologous end joining (NHEJ) and homology-directed repair (HDR). Disruptive NHEJ can occur at any stage throughout the cell cycle in the absence of homologous DNA. In contrast, introduction of specific mutations via HDR is restricted to the S and G2 phases and depends upon the availability of a suitable DNA template. Thus, HDR-based genome editing approaches are often restricted by unwanted NHEJ events, particularly in clinically relevant systems. Overcoming this limitation would greatly increase the therapeutic potential of genome editing.
The student will utilise the Genome Stability Unit’s expertise in DNA repair and know-how in gene editing technology to force HDR and suppress NHEJ during CRISPR-Cas9 gene correction. You will use our EGFP to BFP conversion assay (Glaser A., et al Mol Ther Nucleic Acids 2016 doi: 10.1038/mtna.2016.48) to investigate DNA repair pathway choice, and to identify novel strategies that bias CRISPR/Cas9-mediated genome editing towards HDR.1 This will be achieved through the fusion of protein domains lending HDR-promoting qualities to Cas9.2 Fusions that promote HDR will then be tested in model gene editing systems, for eventual translation into human use – the project here allows some flexibility based on student interest or expertise as to the exact disease to be modelled. This is an exciting project, that has direct translational potential for application in near-future use of gene editing in treatment of human disease.
A/Prof Andrew Deans
Dr Astrid Glaser
Mutations to DNA cause many different types of human disease, including all cancers. DNA repair mechanisms have evolved to protect cells from mutation, and DNA repair genes are therefore important tumour suppressors. The Genome Stability Unit at St Vincent’s Institute researches the mechanisms by which DNA repair is controlled, or could be targeted in therapy. The group currently has a high researcher: student ratio meaning plenty of expert advice is available to students.
For further information about this project, contact: