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United States Department of Agriculture

Agricultural Research Service

Title: Cryopreservation of Shoot Tips of Blackberry and Raspberry by Encapsulation-Dehydration and Vitrification

Authors
item Gupta, Sandhya - PUSA CAMPUS, NEW DELHI I
item Reed, Barbara

Submitted to: CryoLetters
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: December 14, 2005
Publication Date: March 15, 2006
Citation: Gupta, S., Reed, B.M. 2006. Cryopreservation of shoot tips of blackberry and raspberry by encapsulation-dehydration and vitrification. CryoLetters. 27:29-42.

Interpretive Summary: Two procedures were improved for long-term storage of shoot tips of raspberry and blackberry plants in liquid nitrogen (-320 F). Four-week cold treatments of the plant allowed them to be stored. For one protocol shoot tips were enclosed in alginate beads and dried to ~20% moisture content, then plunged in liquid nitrogen. The second technique used chemicals to acclimate the shoots, followed by direct immersion in liquid nitrogen. Shoot tips of 25 types of blackberries and raspberries were successfully cryopreserved in liquid nitrogen with recovery of 60-100% using the encapsulation-dehydration protocol. These results indicate that both of the procedures are promising for cryopreservation of a wide range of Rubus genetic resources.

Technical Abstract: Encapsulation-dehydration and PVS2-vitrification cryopreservation protocols were improved for long-term conservation of a diverse group of Rubus germplasm. Four-week cold acclimation was necessary for recovery of shoot apices of blackberry and raspberry genotypes following cryopreservation. For the encapsulation-dehydration protocol encapsulated apices were pretreated in 0.75 M sucrose for 20 h, dehydrated 6-h under laminar flow to ~20% moisture content on fresh weight basis, then plunged in liquid nitrogen (LN) and rapidly warmed. The PVS2 vitrification protocol included pretreating shoot tips on 5% DMSO medium for 48 h, exposure to loading solution (LS) and PVS2 for 20 min each at 25ºC, followed by immersion in LN and rapid warming. Shoot tips of 25 genotypes in 9 Rubus species were successfully cryopreserved with recovery of 60-100% using the encapsulation-dehydration protocol. Four genotypes of 3 species were tested using the vitrification protocol with 71% average regrowth. These results indicate that both of the protocols optimized in the present study are promising for cryopreservation of a wide range of Rubus genetic resources. Encapsulation-dehydration and PVS2-vitrification cryopreservation protocols were optimized for long-term conservation of a diverse group of Rubus germplasm. Four-week cold acclimation was necessary for recovery of shoot apices of blackberry and raspberry genotypes following cryopreservation. For the encapsulation-dehydration protocol encapsulated apices were pretreated in 0.75 M sucrose for 20 h, dehydrated 6-h under laminar flow to ~20% moisture content on fresh weight basis, then plunged in liquid nitrogen (LN) and rapidly warmed. The PVS2 vitrification protocol included pretreating shoot tips on 5% DMSO medium for 48 h, exposure to loading solution (LS) and PVS2 for 20 min each at 25ºC, followed by immersion in LN and rapid warming. Shoot tips of 25 genotypes in 9 Rubus species were successfully cryopreserved with recovery of 60-100% using the encapsulation-dehydration protocol. Four genotypes of 3 species were tested using the vitrification protocol with 71% average regrowth. These results indicate that both of the protocols optimized in the present study are promising for cryopreservation of a wide range of Rubus genetic resources.

Last Modified: 4/17/2014
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