Phylogenomic analysis by chromosome sorting and painting.

Abstract

Chromosome sorting by flow cytometry is the principle source of chromosome-specific DNA not only for chromosome painting, but also for many other types of genomic analysis such as library construction, discovery and isolation of genes, chromosome specific direct DNA selection, and array painting. Chromosome sorting coupled with chromosome painting is a rapid method for global phylogenomic comparisons. These two techniques have made notable contributions to our knowledge of the evolution of the mammalian genome. The flow sorting of multiple species allows reciprocal painting and permits the delineation of subchromosomal homology and the definition of chromosomal breakpoints. Chromosomes are valuable phylogenetic makers because rearrangements that become fixed at the species level are considered rare events and apparently tightly bound to the speciation process. This chapter covers the preparation of a single chromosome suspension from cell cultures, bivariate chromosome flow sorting, preparation of chromosome paints by degenerate oligonucleotide primed-PCR and the fluorescence in-situ hybridization and detection of whole chromosome specific probes.