Preparation of fluorescent in situ hybridisation probes without the need for optimisation of fragmentation
Patrick J. McCoy,
Anthony J. Costello,
Niall M. Corcoran,
Christopher M. Hovens,
Michael J. Clarkson
Affiliations
Patrick J. McCoy
Departments of Surgery and Urology, University of Melbourne, Royal Melbourne Hospital, Parkville, Australia; Australian Prostate Cancer Research Centre Epworth, Victoria, Australia; Corresponding author at: Departments of Surgery and Urology, University of Melbourne, Royal Melbourne Hospital, Parkville, Australia.
Anthony J. Costello
Departments of Surgery and Urology, University of Melbourne, Royal Melbourne Hospital, Parkville, Australia; Australian Prostate Cancer Research Centre Epworth, Victoria, Australia
Niall M. Corcoran
Departments of Surgery and Urology, University of Melbourne, Royal Melbourne Hospital, Parkville, Australia; Australian Prostate Cancer Research Centre Epworth, Victoria, Australia
Christopher M. Hovens
Departments of Surgery and Urology, University of Melbourne, Royal Melbourne Hospital, Parkville, Australia; Australian Prostate Cancer Research Centre Epworth, Victoria, Australia
Michael J. Clarkson
Departments of Surgery and Urology, University of Melbourne, Royal Melbourne Hospital, Parkville, Australia; Australian Prostate Cancer Research Centre Epworth, Victoria, Australia
DNA-fluorescence in situ hybridisation (DNA-FISH) allows visualisation of chromosome organisation and rearrangement. FISH probes are pools of short fluorescently labelled DNA fragments that are often produced from template plasmids that contain large genomic inserts. For effective sample penetration and target hybridisation it is critical that probe fragments are between 200 and 500bp. Production of these short probes requires significant optimisation and can be confounded access to expensive sonication equipment or inherent sequence features that influence enzymatic fragmentation or amplification. Here we demonstrate that effective FISH probes can be prepared without the need for optimisation of fragmentation using a cocktail of two the 4bp recognition sequence restriction enzymes CviQI and AluI. Method name: Rapid probe construction for Fluorescence in situ hybridization (FISH), Keywords: Fluorescence in situ hybridization, FISH, Cancer, Translocation, Probe
