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| Study on Cryopreservation of XH33 Embryogenic Cells in Gossypium barbadense by Droplet Vitrification |
| LI Lun-Zheng*, GULINAER·Aibaidula*, YANG Rui-Si, ZHOU Jing, CHEN Quan-Jia, QU Yan-Ying, ZHANG Xia** |
| College of Agriculture/Key Laboratory of Agricultural Biotechnology, Xinjiang Agricultural University, Urumqi 830052, China |
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Abstract At present, cotton transgenic technology highly depends on somatic embryogenesis,and preservation of the cultured tissues needs subculture which might cause genetic variation or species degradation of preserved materials so that it is not conducive to the preservation and utilization of resources. Ultra-low temperature preservation can maximize the genetic stability of preserved materials. To establish the method of cryopreservation of Gossypium barbadensee XH33 embryonic cells by droplet vitrification, the present study optimized the system from preculture, vitrification solution and processing time, defrost method, and recovery cultivation, respectively. And the genetic stability of embryonic cell after cryopreservation was checked through the methods of flow cytometry, SSR molecular markers and physiological index detection. The results showed that XH 33 embryogenic cells were pre-cultured in 0.3、0.5、0.7 mol/L sucrose medium in sequence at 4 ℃ for 3 d, and then loaded and dehydrated for 20 min in the modified PVS2 (plant vitrification solutions 2). After stored in liquid nitrogen for 12 h, the cells were thawed in 40 ℃ water bath and washed with 0.5, 0.7, 1.0 mol/L gradient concentration of sucrose solution and then cultured in darkness for 3 d. The cell survival rate was as high as 55.55%. The results of flow cytometry and SSR molecular markers showed that cryopreservation did not affect the genetic stability of cells, and there were no significant changes in the physiological activities of cells after cryopreservation. This study established the cryopreservation system for embryogenic cells of G.barbadensehave by vitrification which provides a reference for long-term preservation of embryogenic cells in island cotton.
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Received: 19 November 2018
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Corresponding Authors:
Xiazhangxjnd@sohu.com
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