Effects of Ascorbic Acid on PVS2 Cryopreservation of Dendrobium Bobby Messina’s PLBs Supported with SEM Analysis

• Published in: Applied biochemistry and biotechnology Vol. 171; no. 2; pp. 315 - 329
• Main Authors: Antony, Jessica Jeyanthi James, Keng, Chan Lai, Mahmood, Maziah, Subramaniam, Sreeramanan
• Format: Journal Article
• Language: English
• Published: Boston Springer US 01.09.2013; Springer; Springer Nature B.V
• Subjects: Acids; Ascorbic Acid - pharmacology; Biochemistry; Biological and medical sciences; Biotechnology; Cell Survival - drug effects; Chemistry; Chemistry and Materials Science; Cryopreservation - methods; Dendrobium - cytology; Dendrobium - drug effects; Dendrobium - growth & development; Dendrobium - ultrastructure; Dose-Response Relationship, Drug; Fundamental and applied biological sciences. Psychology; Growth regulators; Microscopy, Electrochemical, Scanning; Plant biology; Regrowth; Scanning electron microscopy; Temperature; Time Factors; Italy, Sicilia, Messina; Bobby Messina; SEM; Ascorbic acid; Cryopreservation; Regeneration rates; Regeneration; Dendrobium Bobby Messina; Support
• ISSN: 0273-2289; 1559-0291
• DOI: 10.1007/s12010-013-0369-x

Regrowth of the cryopreserved protocorm-like bodies (PLBs) of Dendrobium Bobby Messina was assessed based on the plant vitrification solution 2 (PVS2) optimisation conditions. The optimized protocol obtained based on TTC spectrophotometrical analysis and growth recovery were 3–4 mm of PLBs size precultured in 0.2 M sucrose for 1 day, treated with a mixture of 2 M glycerol and 0.4 M sucrose supplemented with half-strength liquid MS media at 25 °C for 20 min and subsequently dehydrated with PVS2 at 0 °C for 20 min prior to storage in liquid nitrogen. Following rapid warming in a water bath at 40 °C for 90 s, PLBs were treated with unloading solution containing half-strength liquid MS media supplemented with 1.2 M sucrose. Subsequently, the PLBs were cultured on half-strength semi-solid MS media supplemented with 2 % ( w / v ) sucrose without any growth regulators and resulted in 40 % growth recovery. In addition, ascorbic acid treatment was used to evaluate the regeneration process of cryopreserved PLBs. However, growth recovery rates of non-cryopreserved and cryopreserved PLBs were 30 and 10 % when 0.6 mM ascorbic acid was added. Scanning electron microscopy analysis indicates that there are not much damages observed on both cryopreserved and non-cryopreserved PLBs in comparison to PLBs stock culture.