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  • Title: SEED DESICCATION TOLERANCE AND CRYOPRESERVATION OF PHILIPPINE CALAMANSI [x Citrofortunella microcarpa (BUNGE) WIJNANDS].
    Author: Damasco OL, Refuerzo LC.
    Journal: Cryo Letters; 2016; 37(4):219-230. PubMed ID: 27925005.
    Abstract:
    BACKGROUND: The traditional on farm conservation of Calamansi [x Citrofortunella microcarpa (Bunge) Wijnands], an important indigenous Citrus species in the Philippines, is now being threatened by shifting agricultural crop production, climate change, and increasing biotic and abiotic stresses. OBJECTIVE: The study aimed to characterize the desiccation and cryopreservation tolerance of seeds as the basis for complementary long term ex situ conservation. MATERIALS AND METHODS: Intact seeds were desiccated in an airtight container filled with activated silica gel for 0-96 h. Seeds placed in cryotubes were subjected to rapid freezing in liquid nitrogen, rapid thawing in a water bath at 50 degree C for 3 min, and cultured on MS basal medium for seedling recovery and growth. Recovered seedlings were potted out in plastic bags filled with coir dust: garden soil mixture (1:1 v/v) and maintained in the nursery. RESULTS: Significant reduction in percentage germination was obtained at in a moisture content (MC) window between 24.3% and 4.2% and complete loss of viability at below 3.2%. The number of germinated embryos per seed was significantly reduced following desiccation from a mean of 4.2 embryos per seed for the untreated control to 1.2 to 1.02 embryos per seed at 33.3-4.2% MC, respectively. Recovery and germination of seeds after cryopreservation were obtained in a MC window between 24.3% and 4.2% with the maximum seed germination (27%) obtained at 13.4%. Germination abnormalities such as incomplete germination, greening and or enlargement of cotyledon without shoot emergence were observed in both desiccated and cryopreserved seeds. Variations in response to seed desiccation and cryopreservation were observed among Calamansi accessions tested. Maximum seedling recovery after liquid nitrogen storage varied between 12.5% and 61.5%. Recovered seedlings from desiccation and cryopreservation treatments survived ex vitro establishment and showed normal growth and similar morphology with the non-treated control seedlings. CONCLUSION: The partial tolerance of Calamansi seeds to desiccation and subsequent recovery after cryopreservation provides the basis for long term ex situ preservation of this valuable germplasm, although further optimization is needed.
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