Studies on changes in the cellular pattern of cryopreserved explants of Saccharum spp and sugarcane cultivar

Cryopreserved samples in sugarcane generally require a long culturing period and can experience unexpected stress due to ultra-low freezing temperatures. This study aims to investigate the factors that hinder long-term storage by examining the anatomical changes that occur during the cryopreservation of meristem-derived axillary buds in sugarcane. Light microscopy was employed to analyze the microstructure of both cryopreserved and non-cryopreserved samples. The meristem-derived axillary buds from Saccharum spontaneum clones (IND 2019 2045 and IND 2019 2035) and the sugarcane cultivar Co 11015 were cross sectioned at various time intervals: 24 hours, 3 days, 7 days, 14 days, and 21 days after treatment with Liquid Nitrogen (LN) and subsequent rewarming procedures. This analysis aimed to infer the changes in cellular structures due to cryopreservation. The cross-sections of the meristem-derived axillary buds displayed damage to the epidermal and hypodermal regions at different treatment and rewarming durations. Additionally, vascular bundles were found to be highly disorganized and collapsed. The ultra-low temperature freezing likely contributed to ice nucleation within the cells, leading to the destruction of cellular integrity. Among the clones evaluated, Co 11015 demonstrated less damage compared to IND 2019 2045 and IND 2019 2035, suggesting that the commercial cultivar possesses a better capacity to withstand cryoinjury. Furthermore, utilizing analytical tools will enhance our understanding of the harm inflicted on biological tissues by cryopreservation. This knowledge will enable us to refine protocols for the successful regeneration of plantlets, ensuring the long-term conservation of germplasm.