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Evaluation of DNA methods for differentiation of inbred strains of rats and mice

Sharp, Margaret (2001) Evaluation of DNA methods for differentiation of inbred strains of rats and mice. Honours thesis, Murdoch University.

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DNA based technology offers improved differentiating methods in the laboratories of breeders of genetically uniform animals. This project examined four techniques and their application to differentiation of inbred strains of rats and mice.

RAPD-PCR using 10mer primers of arbitrary sequence was evaluated as a method for differentiating strains. Although polymorphisms could be identified, it was not easy to detect these in the complex pattern produced. Attempts to simplify the pattern by reducing the level of dNTP's in the PCR reaction were unsuccessful. Restriction endonuclease digestion (including double digests) of RAPD-PCR products was also evaluated for a range of enzymes. The pattern was not simplified by any of these combinations. However, one endonuclease produced differentiating polymorphisms that were suitable for examination.

Five mouse microsatellites that enabled differentiation between thirteen inbred strains, and six rat microsatellites that enabled differentiation between eight inbred strains and two breeding lines for two outbred strains, were identified using PCR amplification. Multiplexing of two of the rat microsatellites was possible.

Preliminary investigations to adapt Microsatellite Fragment Length Polymorphism techniques for application to differentiation of rats and mice was undertaken. Results indicate the requirement for larger primer anchors in animal work than previously used for non-animal applications.

Short Interspersed Nuclear Element-PCR to detect inter-element length polymorphisms was not successful.

The results of this study showed that, of the techniques evaluated, microsatellite amplification was the cheapest and most reliable method of differentiation of closely related strains of rats and mice.

Item Type: Thesis (Honours)
Murdoch Affiliation(s): School of Biological Sciences and Biotechnology
Notes: Note to the author: If you would like to make your thesis openly available on Murdoch University Library's Research Repository, please contact: Thank you.
Supervisor(s): O'Brien, Philip
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