Automated Quantification of Meiotic Recombination Foci Position and Intensity

During meiosis, homologous chromosomes reciprocally exchange segments of DNA via the formation of crossovers. However, the frequency and position of crossover events along chromosomes are not random. Each chromosome must receive at least one crossover, and the formation of a crossover at one location inhibits the formation of additional crossovers nearby. These crossover patterning phenomena are referred to as "crossover assurance" and "crossover interference," respectively. One key method for quantifying meiotic crossover patterning is to immunocytologically measure the position and intensity of crossover-associated protein foci along the length of meiotic prophase I chromosomes. This approach was recently used to map the position of a conserved E3 ligase, HEI10, along Arabidopsis pachytene chromosomes, providing experimental support for a novel mechanistic "coarsening model" for crossover patterning. Here we describe a user-friendly method for automatically measuring the position and intensity of recombination-associated foci along meiotic prophase I chromosomes that is broadly applicable to studies in different eukaryotic species.