Development of simple storage methods for donor cells such as freeze-drying for bovine somatic cell nuclear tramsfer

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Simple storage methods for donor cells used for somatic cell nuclear transfer were examined. First, we examined whether live cells can be retrieved from simple frozen tissues. Bovine several tissues were frozen at -20 oC, -80 oC and -196 oC. After thawing tissues, live cells were obtained from ear tissues frozen at -80 oC even after 6 months. Next, nuclear status of somatic cells after freeze-drying was examined. Trehalose was introduced into cells taken from ear tissues by digitonin, then the cells were freeze-dried. Fragmentation of nuclear DNA was examined by a Comet assay method. DNA fragmentation of freeze-dried cells treated with 0 or 20g/ml digitonin and 0.4M trehalose was similar to that of untreated live cells.