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United States Department of Agriculture

Agricultural Research Service

Research Project: PRESERVATION AND QUALITY ASSESSMENT OF PLANT GENETIC RESOURCES

Location: Plant And Animal Genetic Resources Preservation Research Unit

Title: Evaluation of cryogenic procedures for cryopreservation of Cassava genotypes

Authors
item Rodrigues, Leonardo -
item Alves, Alfredo -
item Paiva, Luciano -
item Paiva, Renato -
item Jenderek, Maria
item Ellis, David

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: October 16, 2011
Publication Date: October 13, 2011
Citation: Rodrigues, L.A., Alves, A., Paiva, L.V., Paiva, R., Jenderek, M.M., Ellis, D.D. 2011. Evaluation of cryogenic procedures for cryopreservation of Cassava genotypes. Meeting Abstract. Crop Science, San Antonio, TX, October 16-19, 2011.

Interpretive Summary: Cassava (Manihot esculent Crantz) is a perennial plant widely grown in tropical countries and is one of the most important commercial crops globally. Global cassava production in 2009 was 242 million tons. Because of its economic importance to the developing world, conservation efforts to save cassava diversity are underway. Our research looks at the long-term (100+ years) storage of cassava using very cold temperatures in liquid nitrogen (-196oC). Since we need large numbers of uniform shoots for this research, the first phase of the work was to determine what nutrient medium was best for shoot growth. After this was established, 1-2mm shoot tips were excised from of two cassava cultivars, CM 507-37 and COL 1468 and were subsequently treated with a cryoprotectant for varying time periods prior to immersion into liquid nitrogen. Survival following liquid nitrogen was very low and it is theorized that exposure to the cryoprotectant was poor. Subsequent research is focused on providing uniform exposure to PVS2.

Technical Abstract: Cassava (Manihot esculent Crantz) is a perennial plant widely grown in many tropical countries as one of the most important commercial crops. The global cassava production in 2009 was at 242 million tons. Because of its economic importance to a large number of developing world, the application of advanced technology to cassava it is primordial. The cryopreservation in liquid nitrogen (LN2) is the best choice for long-term storage of plant germplasm. In this sense, the cryopreservation of shoot tips of cassava using conventional vitrification method was reported. However, availability and development of simple and effective micropropagation protocol, which can supply a large number of uniform and ideal apices required for successful cryopreservation are the basic requirements. In this study, shoot tips of cassava (Manihot esculenta Crantz cv. CM 507-37 and COL 1468) in vitro plantlets were cryopreserved using the vitrification method. Mononodal microcuttings of 5 mm length with an axillary bud were taken from 2-month-old stock cultures and densely cultured on 2 diferent fresh media (Manteinance x Wild media) in a Petri dish 90 mm × 20 mm. After 14 days, were excised shoot tips of 1 mm from in vitro plantlets and then, placed on solidified pre-culture media, supplemented with 0.3 M sucrose for 16 h; these were placed in growth room in place protect from direct light. The osmo-protected shoot tips were sufficiently dehydrated with a highly concentrated vitrification solution (designated PVS2) for different times prior to plunging into LN2. Successfully vitrified shoot tips were rewarmed rapidly for 1 min in water bath at 45 °C and then plated on culture medium. These vitrified shoot tips developed shoots within 4-6 weeks after being recultured. The results showed that the best culture media, considering the speed and uniformity, was the Wild Cassava media, and that exposure of the shoot tips to PVS2 more than 30 minutes make it impossible to regeneration of this material. Further studies are necessary to adjust a method that can be routinely applied to preserve these lines.

Last Modified: 10/1/2014
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