Use of Uncontrolled Freezing for Liquid Nitrogen Storage of Phytophthora Species. P. W. Tooley, Research Plant Pathologist, U.S. Department of Agriculture, Agricultural Research Service, Foreign Disease-Weed Science Research, Fort Detrick, Bldg. 1301, Frederick, MD 21701. . Plant Dis. 72:680-682. Accepted for publication 24 February 1988. This article is in the public domain and not copyrightable. It may be freely reprinted with customary crediting of the source. The American Phytopathological Society, 1988. DOI: 10.1094/PD-72-0680.

Simple, inexpensive techniques were developed to allow storage of Phytophthora species in liquid nitrogen without the use of a programmable slow-freezing unit. Agar plugs containing mycelium were suspended in 1 ml of cryoprotectant (10% glycerol or 5% dimethyl sulfoxide) in screw-cap polypropylene vials and placed in freezers at –20 C and/or –80 C before storage in liquid nitrogen. High recovery rates (75–100%) were obtained for samples of P. infestans placed at –20 C for 24 min then –80 C for 60 min, and for samples placed at –80 C alone for 30, 45, 60, or 84 min before freezing in liquid nitrogen. Recovery rates of 68–100% were obtained after 7 days when subcultures of 10 different Phytophthora species were placed at –80 C for 60 min before storage in liquid nitrogen. Isolates of P. infestans, P. boehmeriae, and P. megasperma f. sp. glycinea frozen by this method showed no decrease in viability following at least 9 mo of storage in liquid nitrogen. The success of this technique may allow more widespread use of liquid nitrogen storage technology for Phytophthora spp.