Abstract Chromosome painting (CP) refers to fluorescence in situ hybridization (FISH) of chromosome‐specific DNA probes to identify large chromosome regions, chromosome arms, and whole chromosomes. For CP and CCP (comparative chromosome painting) in plants, most often, contigs of chromosome‐specific bacterial artificial chromosomes (BAC) from the species of origin or a related species are used as painting probes. CP enables visualization and tracing of particular chromosome regions and/or chromosomes throughout all mitotic and meiotic stages as well as the corresponding interphase chromosome territories. CCP enables identification of large‐scale homeologous chromosome regions and chromosomes shared among two or more species. Here, a step‐by‐step protocol for carrying out CP in Arabidopsis thaliana (Arabidopsis) and CCP in other crucifer taxa based on the use of Arabidopsis chromosome‐specific BAC contigs is described. © 2016 by John Wiley & Sons, Inc.