Identification of germline competent chimaeras by copulatory plug genotyping |
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Authors: | Stephen Wilson Steven A Sheardown |
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Institution: | (1) Molecular Discovery Research, GlaxoSmithKline Pharmaceuticals, New Frontiers Science Park, Harlow, Essex, CM19 5AD, UK; |
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Abstract: | The increasing use of ES cell lines from strains other than 129, in particular C57BL/6, has greatly reduced the time taken
to generate gene knockouts on a defined genetic background. Generally, C57BL/6 ES cell lines transmit less efficiently through
the germline than 129 lines; consequently the burden on animal husbandry at this stage is increased. Genotyping sperm from
chimaeric males may be used to identify mice which are transmitting the manipulated allele, however it requires that the mice
be culled and the sperm used for IVF. Here we describe a quick and reliable method for genotyping copulatory plugs. Males
which produce a positive result can then be naturally mated. Thus far we have observed a perfect correlation between copulatory
plug genotype and germline transmission, accompanied by considerable savings in mouse numbers and resources. |
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