Protocol

Biological Aging

Volume 371 of the series Methods in Molecular Biology™ pp 267-287

Detection and Analysis of Somatic Mutations at a lacZ Reporter Locus in Higher Organisms

Application to Mus musculus and Drosophila melanogaster
  • Ana Maria GarciaAffiliated withUniversity of Texas at San Antonio
  • , Rita A. BusuttilAffiliated withBuck Institute for Age Research
  • , Armando RodriguezAffiliated withUniversity of Texas Health Sciences Center
  • , Carlos CabreraAffiliated withUniversity of Texas Health Sciences Center
  • , Martha LundellAffiliated withUniversity of Texas at San Antonio
  • , Martijn E. T. DolléAffiliated withNational Institute of Public Health and Environment
  • , Jan VijgAffiliated withBuck Institute for Age Research

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Abstract

Methods to detect and analyze somatic mutations in higher organisms are critically important in view of their causal role in cancer, heritable diseases, and, possibly, aging. Here, we describe detailed protocols for the use of a mutational reporter system based on lacZ-containing plasmids integrated in the germline of Mus musculus and Drosophila melanogaster. Plasmids containing the bacterial lacZ gene integrated at one or more chromosomal sites can be excised, purified and recovered in suitable Escherichia coli hosts allowing the positive selection of mutant lacZ genes and their further molecular characterization. This system is capable of detecting a broad range of mutational events, varying from small mutations in the lacZ reporter gene to large genome rearrangements with one breakpoint in lacZ and the other breakpoint elsewhere in the genome.

Key Words

Somatic mutations genome rearrangements lacZ-plasmids Mus musculus Drosophila melanogaster