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Time-lapse tracking of barley androgenesis reveals position-determined cell death within pro-embryos

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Author: Maraschin, S.D.F. · Vennik, M. · Lamers, G.E.M. · Spaink, H.P. · Wang, M.
Source:Planta, 4, 220, 531-540
Identifier: 238321
doi: doi:10.1007/s00425-004-1371-x
Keywords: Androgenesis · Microspore embryogenesis · Time-lapse tracking · Cell membranes · Morphology · Starch · Stress analysis · Abiotic stress · Embryogenic potential · Microspores · Time-lapse tracking · Crops · bacterial spore · barley · cell death · cell survival · cytology · electron microscopy · genetics · growth, development and aging · haploidy · microscopy · physiology · plant seed · reproduction · ultrastructure · Cell Death · Cell Survival · Haploidy · Hordeum · Microscopy, Electron · Microscopy, Video · Reproduction · Seeds · Spores · Anatomy · Cell Walls · Farm Crops · Starch · Stresses · Hordeum · Hordeum vulgare subsp. vulgare


Following abiotic stress to induce barley (Hordeum vulgare L.) androgenesis, the development of 794 enlarged microspores in culture was monitored by time-lapse tracking. In total, 11% of the microspores tracked developed into embryo-like structures (type-I pathway), 36% formed multicellular structures (type-II pathway) and 53% of the microspores followed gametophytic divisions, accumulated starch and died in the first days of tracking (type-III pathway). Despite the microspore fate, enlarged microspores showed similar morphologies directly after stress treatment. Ultrastructural analysis, however, revealed two morphologically distinct cell types. Cells with a thin intine layer and an undifferentiated cytoplasm after stress treatment were associated with type-I and type-II pathways, whereas the presence of differentiated amyloplasts and a thick inline layer were associated with the type-III pathway. Tracking revealed that the first morphological change associated with embryogenic potential was a star-like morphology, which was a transitory stage between uninucleate vacuolated microspores after stress and the initiation of cell division. The difference between type-I and type-II pathways was observed during the time they displayed the star-like morphology. During the transition phase, embryo-like structures in the type-I pathway were always released out of the exine wall at the opposite side of the pollen germ pore, whereas in the type-II pathway multicellular structures were unable to break the exine and to release embryo-like structures. Moreover, by combining viability studies with cell tracking, we show that release of embryo-like structures was preceded by a decrease in viability of the cells positioned at the site of exine wall rupture. These cells were also positively stained by Sytox orange, a cell death indicator. Thereby, we demonstrate, for the first time, that a position-determined cell death process marks the transition from a multicellular structure into an embryo-like structure during barley androgenesis. © Springer-Verlag 2004.