Gynoecium pubescence in soybean: a prevalent false-positive during in vitro androgenesis

Doubled haploidy serves as an effective tool for enhancing genetic gain in plant breeding programs. Androgenesis, an extensively-employed form of doubled haploidy, involves totipotent induction within the immature microgametophyte followed by agamous embryogenesis and chromosome doubling. Protocol development for in vitro androgenesis is reliant upon the application of species- (and oftentimes genotype-) specific induction stimuli as well as phenotypic markers that reflect cellular reprogramming. In cell cultures, however, the latter is impeded readily by pseudo-embryogenic structures of somatic origin. In this study, gynoecium-derived trichomes of soybean were intentionally incorporated into isolated microspore cultures. Trichome morphology was then compared to early-dividing zygotic and apozygotic plant embryos previously reported, as were nuclei orientation and cross wall composition. Lastly, autofluorescence intensity was measured for trichomes and various microspore phenotypes, the results of which implied that autofluorescence post-light exposure may be a useful parameter for culture purification and/or quality control. The findings herein demonstrate that trichomes serve a pseudo-embryogenic role during in vitro androgenesis in soybean and necessitate extra caution when identifying sporophytic tissues in species with floral pubescence.