Imaging chromosome dynamics in meiosis in plants.

Progression of meiosis has been traditionally reconstructed from microscopic images collected from fixed cells. This approach has clear shortcomings in accurately portraying the dynamics of meiotic processes. Studies conducted in recent years mostly in unicellular fungi have shown that chromosomes in meiotic prophase exhibit dynamic motility that cannot be accurately examined using fixed cell imaging. However, in contrast to yeast, research on meiotic chromosome dynamics in multicellular eukaryotes has been lagging. This was in part because meiocytes in multicellular eukaryotes reside deep within reproductive organs and are often refractory to culturing. Here, we describe a method in which intact, live-plant reproductive organs (anthers) are cultured to enable monitoring chromosome dynamics of meiocytes using multiphoton excitation (MPE) microscopy. The method was developed for use in maize but can be applied to other plant species and adapted for use in other taxa in which meiocytes are embedded in multicellular reproductive structures. MPE microscopy allows visualization of meiocytes embedded within native tissue in planta and thus meiocytes remain intact for the entire imaging procedure. We detail the kinds of time-lapse movies that can be captured and analyzed using this technique and also highlight software packages that can be utilized for analysis of movies chromosome dynamic in live meiocytes.