ISOLATION OF HIGH-MOLECULAR-WEIGHT DNA FOR RELIABLE GENOTYPING OF TRANSGENIC MICE

A fast and reliable method for the PCR characterization of DNA from mo use toes is described. The toes biopsied to rag the mice are incubated for 2 h in proteinase K and heated for 15 min at 95 degrees C. This D NA solution is directly used as a template for PCR amplification. The same procedure can be used for PCR analysis of DNA from other tissues in adult mice, mouse embryos and cultured cells. Because of minimal ti ssue manipulation, high-quality and high-molecular-weight DNA (fragmen ts larger than 100-200 kb) is isolated. This procedure is performed in a single tube and requires no organic solvent extraction or centrifug ation, allowing the isolation of high-molecular-weight DNA suitable fo r PCR amplification in a fast and reproducible way. Only tile tissue e xcised during mice tagging is used and a large number of animals can h e qucikly and simultaneously analyzed as required to maintain a transg enic mice colony. In addition, this rapid and efficient procedure repr esents an alternative to other methods in which, in our experience, in hibition of the PCR amplification occurs when DNA from rail tissues is used.