Development of an Efficient Genotyping Method to Detect Obese Mutation in the  Mouse Leptin Gene for Use in SPF Barrier Facilities

Ayabe H, Ikeda S, Maruyama S, Shioyama S, Kikuchi M, Kawaguchi A, Yamada T, Ikeda T.

We have developed a rapid and efficient genotyping method for detection of the mouse leptin obese mutation (Lep(ob)) using tetra-primer amplification refractory mutation system-polymerase chain reaction (tetra-primer ARMS-PCR). In this method, whole blood collected onto gamma-ray sterilized Flinders Technology Associates (FTA) filter paper is used as PCR template without a DNA purification step. Three genotypes (Lep(ob)/Lep(ob), Lep(ob)/+ and +/+) differentiated by single-tube PCR and electrophoresis were perfectly consistent with those determined by PCR-restriction fragment length polymorphism (PCR-RFLP). This method can save material costs and operation time, because it does not require restriction enzyme digestion and could be set up in most specific pathogen-free (SPF) barrier facilities.

Journal of  Veterinary Medicine Science

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