A reliable technique for karyotyping mouse oocytes prepared by a gradual fixation/air-drying method followed by multicolour FISH

Abstract

Chromosome segregation errors during oocyte meiosis increase with age and lead to aneuploidy; hence, the mechanism has been studied extensively. The mouse is the most widely used experimental animal for this purpose. However, the lack of a reliable and efficient technique for karyotyping mouse oocytes has limited comprehensive studies of chromosome-specific segregation errors in this animal model. Here, we developed a novel karyotyping technique for mouse oocytes by applying multicolour fluorescence in situ hybridisation (FISH) to chromosome slides prepared by a gradual fixation/air-drying method, which is best suited to avoid rupture of oocyte membrane and artificial loss of chromosomes. The success rate of karyotyping meiosis I and II oocytes was about 30%, which improved to over 90% when the oocytes were ‘flattened’ during fixation and the chromosome specimens were denatured at 4°C. When this technique was applied to the karyotyping of meiosis II oocytes from aged female mice and from young female mice injected with colchicine, more than 80% of the oocytes were successfully karyotyped and the number of chromosomes was identified on all aberrant chromosomes. In conclusion, our technique allows for the efficient and reliable karyotyping of mouse oocytes.