Genetic stability of cryopreserved shoot tips of <Emphasis Type="Italic">Rubus</Emphasis> germplasm |
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Authors: | Nina Rosa F Castillo Nahla V Bassil Sugae Wada Barbara M Reed |
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Institution: | (1) Department of Horticulture, Oregon State University, Corvallis, OR, USA;(2) USDA-ARS National Clonal Germplasm Repository, 33447 Peoria Road, Corvallis, OR 97333-2521, USA; |
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Abstract: | Questions often arise concerning the genetic stability of plant materials stored in liquid nitrogen for long time periods.
This study examined the genetic stability of cryopreserved shoot tips of Rubus germplasm that were stored in liquid nitrogen for more than 12 yr, then rewarmed and regrown. We analyzed the genetic stability
of Rubus grabowskii, two blackberry cultivars (“Hillemeyer” and ‘Silvan’), and one raspberry cultivar (“Mandarin”) as in vitro shoots and as field-grown plants. No morphological differences were observed in greenhouse-grown cryopreserved plants when
compared to the control mother plants. In the field, cryopreserved plants appeared similar but were more vigorous than mother
plants, with larger leaves, fruit, and seeds. Single sequence repeats (SSR) and amplified fragment length polymorphism (AFLP)
analyses were performed on shoots immediately after recovery from cryopreservation and on shoots subcultured for 7 mo before
analysis. Ten SSR primers developed from “Marion” and “Meeker” microsatellite-enriched libraries amplified one to 15 alleles
per locus, with an average of seven alleles and a total of 70 alleles in the four genotypes tested. No SSR polymorphisms were
observed between cryopreserved shoots and the corresponding mother plants regardless of subculture. Although no polymorphisms
were detected in shoots analyzed immediately after recovery from cryopreservation, AFLP polymorphisms were detected in three
of the four Rubus genotypes after they were subcultured for 7 mo. Field-grown plants from the polymorphic shoot tips of R. grabowskii and ‘Silvan’ displayed the same AFLP fingerprints as their corresponding mother plants. Only long-cultured in vitro shoot tips displayed polymorphisms in vitro, and they were no longer detected when the plants were grown ex vitro. The transitory nature of these polymorphisms should be carefully considered when monitoring for genetic stability. |
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