A Method for Detecting Parthenogenesis in Asteraceae Species

Embryogenesis in plants is a process that not only occurs after fertilization but can also be initiated in its absence. This form of embryogenesis is known as parthenogenesis, and it is an essential part of a process called apomixis. Apomixis has long been seen as the holy grail of plant breeding because it produces clonal seeds. Therefore, engineering (single components of) apomixis remains a highly desired research goal. Over the last few years, significant progress has been made toward understanding and engineering both complete synthetic apomixis system and individual components of apomixis, such as parthenogenesis, in crops. Characterization of candidate parthenogenesis genes involves thorough phenotypic analysis and microscopic observation of developing embryos, which are, however, not readily accessible because they are embedded in several integumental layers comprising the protective seed coat. Here, we provide a detailed approach for detecting parthenogenesis and we further demonstrate its application to a sexual crop species, i.e., lettuce. We describe an emasculation technique by decapitation, embryo sac isolation, imaging of cleared embryos, and ploidy analysis of embryo sacs utilizing flow cytometry.