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Title: Droplet-vitrification and morphohistological studies of cryopreserved shoot tips of cultivated and wild pineapple genotypes

Author
item SOUZA, FERNANDA - Labex - Embrapa
item ERGUN, KAYA - Hacettepe University, Turkey
item VIERIA DE JESUS, LIVIA - Labex - Embrapa
item DE SOUZA, EVERTON - Labex - Embrapa
item AMORIM, VANUSIA, B DE O - Labex - Embrapa
item Skogerboe, Dianne
item Matsumoto Brower, Tracie
item ALVES, ALFREDO - Embrapa
item LEDO, CARLOS - Labex - Embrapa
item Jenderek, Maria

Submitted to: Plant Cell Tissue and Organ Culture
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/27/2015
Publication Date: 10/31/2015
Citation: Souza, F.V., Ergun, K., Vieria De Jesus, L., De Souza, E.H., Amorim, V., Skogerboe, D.M., Matsumoto Brower, T.K., Alves, A.A., Ledo, C., Jenderek, M.M. 2015. Droplet-vitrification and morphohistological studies of cryopreserved shoot tips of cultivated and wild pineapple genotypes. Plant Cell Tissue and Organ Culture. doi:10.1007/s11240-015-0899-8.

Interpretive Summary: Pineapple is one of the most economically exploited tropical fruits in the world and in the last few decades, several species of pineapple have been used as material in industrial manufacturing, pharmaceuticals and cosmetics. Pineapple genetic resources are threatened by many abiotical and biotical factors; hence, their conservation is critical to securing their availability for future uses. Conservation in ultra-low temperature (cryopreservation) is considered as the most economic and safe way of maintaining plant material for a long-term. Our study established cryoprocessing factors that produced high plant recovery of diverse cultivated and wild pineapple varieties, after exposure to ultra-low temperatures and described injuries caused by the low temperature in pineapple tissues on a microscopic level. This is the first report on application of a droplet-vitrification technique in that plant category and it will simplify long-term preservation of pineapple germplasm in the U.S and global gene banks.

Technical Abstract: Germplasm conservation of pineapple [Ananas comosus (L.) Mer.] is crucial to preserve the genus’ genetic diversity to secure material for genetic improvement and to support innovative and new research. Long-term conservation is accomplished through cryopreservation that is done by storing cells or tissues at ultra-low temperature in liquid nitrogen (-196 °C). Droplet-vitrification, a combination of droplet freezing and solution-based vitrification, was used to establish a protocol for cryopreservation of pineapple genetic resources. This protocol was tested on cultivated and wild pineapple genotypes to establish a long-term germplasm security duplicate as well as to investigate the cryo-injuries in the tissues by means of histological techniques. Excised shoot tips (0.5 - 1 mm with one primordial leaf) of different pineapple genotypes were precultured for 48 h on solid MS medium containing 0.3 M of sucrose. Three PVS2 exposure times (30, 45 and 60 min) were tested. The results showed high post cryopreservation survival for all genotypes tested. The best PVS2 exposure time varied according to genotype, although 45 min gave the best survival for the majority of genotypes. The technique was highly efficient in cryopreserving meristem shoot tips of different pineapple genotypes, and was also less laborious than techniques previously reported. This is a first report on application of the droplet-vitrification technique to diverse genotypes of cultivated and wild pineapples and the first report on histological changes occurring in cryopreserved Ananas tissue.