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ARS Home » Pacific West Area » Corvallis, Oregon » National Clonal Germplasm Repository » Research » Publications at this Location » Publication #222021
Title: In Vitro Antioxidant Treatment Improves the Recovery of Cryopreserved Blackberry Shoot Tips

Author
item UCHENDU, ESTHER - OREGON STATE UNIVERSITY
item Reed, Barbara

Submitted to: In Vitro Cellular and Developmental Biology
Publication Type: Abstract Only
Publication Acceptance Date: 4/15/2008
Publication Date: 6/12/2008
Citation: Uchendu, E., Reed, B.M. 2008. In Vitro Antioxidant Treatment Improves the Recovery of Cryopreserved Blackberry Shoot Tips. In Vitro Cellular and Developmental Biology. 44:2029.

Interpretive Summary: At the USDA ARS genebank in Corvallis, Oregon, we are interested in the long-term preservation of blackberry plants. We are looking for ways to improve storage in liquid nitrogen (cryopreservation). The storage of plant tissues in liquid nitrogen imposes stress on plant cells and tissues resulting in increased oxidation and damage. This study evaluated the effect of the antioxidant compounds alpha tocopherol (vitamin E) and ascorbic acid (vitamin C) added at critical points in the cryopreservation protocol. Shoot tips of tissue cultured blackberries (1 mm) were cryopreserved with a commonly used protocol. Four points in the protocol were used to test the antioxidants. Vitamin E added at two points greatly improved recovery, resulting in a 60% increase compared to the controls. Vitamin C treatments significantly improved shoot tip regrowth when added at three points in the protocol and increased recovery by 50-60%. Vitamin C was only beneficial during recovery from cryopreservation if iron was not present in the medium. This study indicates that oxidation is an important factor in losses from cryopreservation and that the addition of antioxidants at critical points of the protocol can greatly improve regrowth following cryopreservation.

Technical Abstract: Cryopreservation imposes stress on plant cells and tissues resulting in increased oxidation and damage. This study evaluated the effect of the antioxidant compounds alpha tocopherol (vitamin E) and ascorbic acid (AA) added at critical points in the cryopreservation protocol. Shoot tips of in-vitro grown Rubus (blackberry) cultivars (0.8 -1 mm) were cryopreserved using the PVS2-vitrification technique. Pretreatment of shoot tips for 48 hr on agar plates with 5% dimethyl sulfoxide and 15 mM vitamin E significantly (P<0.0001) improved recovery, resulting in 58% regrowth compared to 35% for the controls. Vitamin E at 20 mM in the post-cryopreservation rinsing solution significantly increased recovery from 45% for controls to 72%. There were no significant effects of vitamin E in the loading solution or the recovery medium. Treatments of 50 to 100 mM AA significantly improved regrowth when added to the loading solution (control 47%, AA 77-88%) or the rinsing solution (control 47%, AA 82-83%). In the standard recovery medium AA was detrimental to regrowth of shoot tips. However, on regrowth medium without iron, regrowth of shoot tips increased from 51% for controls to 85% with 50 mM AA. This study indicates that oxidation is an important factor in losses from cryopreservation and that the addition of antioxidants at critical points of the protocol can greatly improve regrowth following cryopreservation.