Towards Long-Term Storage Of Wheat Pollen
Daniela Impe, Daniel Ballesteros, Till Ischebeck, Claudia Köpnick, Hardy Rolletschek, Michael Melzer, Manuela Nagel, Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Germany, Royal Botanic Gardens, Kew, United Kingdom, Department of Plant Biochemistry, Georg-August University Göttingen, Germany
Cryopreservation of pollen is used to preserve and exchange nuclear genes of plant genetic resources and to facilitate crosses in breeding programs. For many species, pollen preservation protocols have been established. In wheat breeding programs, long- term pollen preservation would be of high interest to improve the crossing management in hybrid production. However, favorable storage conditions for wheat pollen have not been studied yet and physiological, biochemical and genetic processes in pollen after shedding are hardly understood.
The present project aims to investigate factors that influence wheat pollen viability in order to extend its longevity by cryopreservation. Firstly, a viability test and evaluation scheme based on pollen tube growth in vitro was established and comparisons between in vivo germination, staining methods and impedance flow cytometry showed that more than one test is required to determine wheat pollen viability.
Secondly, storage of wheat pollen under four different non-cryogenic conditions showed that wheat pollen lose germination ability rapidly. At room temperature, viability was lost within one hour at both low and high relative humidity (RH). Under cold temperatures (6-8 °C, both RH) pollen survival was extended moderately. Sugar contents (fructose, glucose and sucrose) increased two- to threefold in pollen stored for one hour under all non-cryogenic conditions. Therefore, we speculate that sugars are actively mobilized during pollen storage. To gain a comprehensive overview of processes contributing to viability loss, a metabolomics and transcriptomic study has been initiated and is complimented by transmission electron microscopy that illustrates ultrastructural differences between fresh, stored and dead wheat pollen.
Finally, storage under ultra-low temperatures (- 196°C) is being investigated in detail. A self-made flash drier was used to control the pollen water content by fast drying. The effects of drying and cryogenic storage are investigated using differential scanning calorimetry and will indicate the feasibility of long-term pollen cryopreservation.