Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
8464154 | Cryobiology | 2018 | 4 Pages |
Abstract
Freeze-drying (FD) technique has been applied as an alternative technology to preserve gene resources to allow simple sperm preservation and shipment at 4â¯Â°C. Nevertheless, DNA sperm might be damaged by mechanical or oxidative stress throughout FD procedure. Therefore, suitable protection to maintain DNA integrity is required. The aim of this study was to determine the effect of rosmarinic acid (RA) as an antioxidant and two chelating agents (EGTA and EDTA) on the DNA integrity of freeze-dried rabbit sperm after storage of the samples at 4â¯Â°C and room temperature for 8 months. Rabbit sperm were freeze-dried in basic medium (10â¯mM Tris-HCl buffer and 50â¯mM NaCl) supplemented with 50â¯mM EGTA (1), 50â¯mM EGTA plus 105â¯Î¼M RA (2), 50â¯mM EDTA (3) or 50â¯mM EDTA plus 105â¯Î¼M RA (4). Semen samples were kept at 4â¯Â°C and room temperature during 8 months. After rehydration, DNA integrity was evaluated with Sperm Chromatin Dispersion test observing that DNA fragmentation was higher when semen samples were freeze-dried with EGTA (10.9%) than with EDTA (4.1%) (pâ¯<â¯0.01). Furthermore, RA acted better under adverse conditions and no significant differences were found in temperature storage. Summarizing, FD is a method that can allow simple gene resources preservation among 4â¯Â°C to 25â¯Â°C during 8 months and transportation without the need for liquid nitrogen or dry ice. EDTA chelating agent is the most suitable media for freeze-dried rabbit sperm and the addition of RA protects the DNA against the oxidative stress caused during FD procedure.
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Authors
Paula Domingo, Maite Olaciregui, Noelia González, Ignacio De Blas, Lydia Gil,