A rapid PCR-based method for the identification of ob mutant mice

  • Ellett J
  • Evans Z
  • Zhang G
 et al. 
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Abstract

With increasing incidence of obesity, there is greater demand for suitable research and therapeutic models. The ob/ob mouse model develops obesity by 5 weeks of age. Previously, a method using DNA purification, PCR, and restriction digestion of products was devised to identify mice bearing the ob allele. Here, we describe a direct PCR method that requires no DNA purification. Wild-type and ob-specific primers are used under the same conditions in two separate and simultaneously run three-primer PCRs. Standard PCR using the wild-type primer mix produces 191 bp and 104 bp bands in +/+ and ob/+ and only the control 191 bp band in ob/ob animals. The ob-specific primer reaction produces 191 bp and 123 bp bands in ob/+ and ob/ob and only the control 191 bp band in +/+ animals. Phenotypic weight gain in offspring of heterozygous intercrosses was used to validate genotypes. This primer-specific PCR method allows simultaneous identification of +/+, ob/+, and ob/ob genotypes prior to breeding age to facilitate breeding and research studies in an important model of clinical obesity.

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Authors

  • Justin D. Ellett

  • Zachary P. Evans

  • Guojing Zhang

  • Kenneth D. Chavin

  • Demetri D. Spyropoulos

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