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Open Access Methodology article

Combining M-FISH and Quantum Dot technology for fast chromosomal assignment of transgenic insertions

Mohammed Yusuf1, David LV Bauer1, Daniel M Lipinski2, Robert E MacLaren2, Richard Wade-Martins3, Kalim U Mir1 and Emanuela V Volpi1*

Author Affiliations

1 Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford, OX3 7BN, UK

2 Nuffield Laboratory of Ophthalmology and Oxford Eye Hospital Biomedical Research Centre, University of Oxford, John Radcliffe Hospital, Oxford, OX3 9DU, UK

3 Department of Physiology, Anatomy and Genetics, Oxford Parkinson's Disease Centre, University of Oxford, South Parks Road, Oxford, OX1 3QX, UK

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BMC Biotechnology 2011, 11:121  doi:10.1186/1472-6750-11-121

Published: 13 December 2011

Abstract

Background

Physical mapping of transgenic insertions by Fluorescence in situ Hybridization (FISH) is a reliable and cost-effective technique. Chromosomal assignment is commonly achieved either by concurrent G-banding or by a multi-color FISH approach consisting of iteratively co-hybridizing the transgenic sequence of interest with one or more chromosome-specific probes at a time, until the location of the transgenic insertion is identified.

Results

Here we report a technical development for fast chromosomal assignment of transgenic insertions at the single cell level in mouse and rat models. This comprises a simplified 'single denaturation mixed hybridization' procedure that combines multi-color karyotyping by Multiplex FISH (M-FISH), for simultaneous and unambiguous identification of all chromosomes at once, and the use of a Quantum Dot (QD) conjugate for the transgene detection.

Conclusions

Although the exploitation of the unique optical properties of QD nanocrystals, such as photo-stability and brightness, to improve FISH performance generally has been previously investigated, to our knowledge this is the first report of a purpose-designed molecular cytogenetic protocol in which the combined use of QDs and standard organic fluorophores is specifically tailored to assist gene transfer technology.