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Preserving the Female Genome in Trehalose Glass at Supra-Zero Temperatures: The Relationship Between Moisture Content and DNA Damage in Feline Germinal Vesicles

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Abstract

Introduction

Maintaining a stable dry state is critical for long-term preservation of live biomaterials at suprazero temperatures. The objective of the study was to characterize the effect of moisture content on DNA integrity within the germinal vesicle (GV) of feline oocytes following dehydration and storage at 22–24 °C.

Methods

Using microwave-assisted drying, conditions that led to a predictable and stable moisture content in trehalose solutions were determined. To explore moisture content stability during storage, trehalose samples were dried for 15 min and stored in glass vials at 11 or 43% RH for 8 weeks. To examine whether this condition allowed proper storage of GVs, permeabilized cat oocytes were incubated in trehalose for 10 min and dried for 15 or 30 min. Oocytes then were rehydrated to assess DNA integrity either directly after drying or after 8 weeks of storage in an 11% RH environment. Raman spectroscopy was used to identify the states of dried samples during storage.

Results

Moisture content was stable during the storage period. There was no significant difference in DNA integrity between fresh and dried samples without storage. After 8 weeks of storage, DNA integrity was maintained in GVs dried for 30 min. Samples dried for 15 min and stored were compromised, suggesting crystallization of the preservation matrix during storage. Biostabilization was optimal when samples were directly processed to moisture contents consistent with storage in the glassy state.

Conclusion

Microwave-assisted drying processing and storage conditions were optimized to ensure stable long-term storage of structural and functional properties of genetic resources.

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Acknowledgments

Research reported in this publication was supported by the Office of the Director, National Institutes Of Health under Award Number R01OD023139. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. The authors are indebted to Dr. Alida Kinney and staff at the Cabarrus Spay and Neuter Clinic for generously facilitating the transfer of resources from consenting pet-owners to UNC Charlotte. We also thank Sidney Richards for technical assistance.

Conflict of interest

Shangping Wang, Pei-Chih Lee, Amanda Elsayed, Fan Zhang, Yong Zhang, Pierre Comizzoli, and Gloria D. Elliott declare that they have no conflicts of interest.

Ethical Approval

The study did not require the approval of an Animal Care and Use Committee because cat ovaries were collected at local veterinary clinics as byproducts from owner-requested routine ovario-hysterectomies.

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Authors and Affiliations

  1. Department of Mechanical Engineering and Engineering Sciences, University of North Carolina at Charlotte, 9201 University City Blvd., Charlotte, NC, 28223, USA

    Shangping Wang, Amanda Elsayed & Gloria D. Elliott

  2. Smithsonian Conservation Biology Institute, National Zoological Park, Washington, DC, USA

    Pei-Chih Lee & Pierre Comizzoli

  3. Department of Electrical and Computer Engineering, University of North Carolina at Charlotte, Charlotte, NC, USA

    Fan Zhang & Yong Zhang

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  1. Shangping Wang
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Correspondence to Gloria D. Elliott.

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Wang, S., Lee, PC., Elsayed, A. et al. Preserving the Female Genome in Trehalose Glass at Supra-Zero Temperatures: The Relationship Between Moisture Content and DNA Damage in Feline Germinal Vesicles. Cel. Mol. Bioeng. 14, 101–112 (2021). https://doi.org/10.1007/s12195-020-00635-y

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