Cryopreservation of citrus for long-term conservation

More than 850 varieties of Citrus, Fortunella, and Citrus-related species are maintained within the USDA-ARSNational Plant Germplasm System and the University of California Citrus Variety Collection. These geneticresources are held within duplicated field, screenhouse, and greenhouse collections. Plant materials are atrisk of disease infestations, abiotic stresses, and natural disasters. We have developed a cryopreservationmethod by which we can back-up citrus genetic resources for the long term at liquid nitrogen temperatures. We excise shoot tips directly from greenhouse or screenhouse source plants, surface sterilize, and then treatwith cryoprotectants. These solutions dehydrate and allow the shoot tips to survive liquid nitrogen exposure. Plants are recovered by micrografting thawed shoot tips onto ‘Carrizo’ seedling rootstocks. Plants representingdiverse species exhibit a high regrowth (more than 50%), and can be transferred to greenhouse conditionswithin months. Cryopreserved plants did not revert to a juvenile state. Accessions of Citrus aurantifolia, Citrusclementina, Citrus celebica, and Citrus paradisi all flowered within 13 to 15 months after cryoexposure. Thesemethods are also applicable for disease eradication using cryotherapy techniques.