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ABSTRACT ARCHIVE |
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CryoLetters is a bimonthly, international journal for low temperature science and technology |
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CryoLetters 31 (1), 1-13 (2010) DETECTION OF 8-HYDROXY-2’-DEOXYGUANOSINE AS A MARKER OF OXIDATIVE DAMAGE IN DNA AND GERMPLASM EXPOSED TO CRYOGENIC TREATMENTS Jason W. Johnston1,2*, Isobel Pimbley1, Keith Harding3 and Erica E. Benson3 1University of Abertay Dundee, Bell Street, Dundee DD1 1HG, Scotland, UK. Abstract An HPLC method has been optimised to measure 8-hydroxy-2’-deoxyguanosine (8OHdG) in DNA and germplasm with the objective of using the adduct as a marker of cryostorage stability. The encapsulation-dehydration cryopreservation protocol was adapted as a model system for assessing the formation of 8OHdG from alginate-encapsulated DNA of calf thymus (CT) and currant species (Ribes) exposed to temperatures of -20 and -196şC. The presence of H2O2 exacerbated the formation of 8OHdG in encapsulated CT and Ribes DNA. Production of the oxidized adduct was lower in the plant system. A reduction in residual water following osmotic dehydration and evaporative desiccation was associated with reduced adduct formation in encapsulated DNA. No significant differences in 8OHdG adduct formation were observed in plants regenerated from cryopreserved Ribes meristems derived from genotypes known to have differential tolerance to cryopreservation. Keywords: cryobiology, DNA, free radicals, oxidative stress, 8OHdG, stability
CryoLetters 31 (1), 14-23 (2010) CRYOPRESERVATION OF Thymus moroderi BY DROPLET VITRIFICATION Ana Marco-Medina1, José Luis Casas1*, Rony Swennen2 and Bart Panis2 1Laboratory of Plant Biotechnology, Institute of Biodiversity CIBIO (Centro Iberoamericano de la Biodiversidad), University of Alicant, Crta. San Vicente del Raspeig s/n. E-03690 San
Vicente del Raspeig, Alicante (Spain). Abstract Thymus moroderi Pau ex Martínez (Labiatae) was successfully cryopreserved using the droplet vitrification method. After 20 min in loading solution at room temperature, shoot tips were dehydrated with PVS2 at 0şC for 30 min and immersed into LN. For thawing, shoot-tips were transferred into recovery solution for 15 min. A test of different recovery media revealed that the best results were obtained when the medium was supplement with 0.275 µM BA. Keywords: cryopreservation, droplet vitrification, hyperhydration, in vitro conservation, Thymus moroderi
CryoLetters 31 (1), 24-28 (2010) VITRIFICATION OF MOSSES: A USEFUL METHOD FOR THE CRYOPRESERVATION OF Splachnum ampullaceum Hedw. Rubén Mallón1,3, Juan Rodríguez-Oubińa2 and María Luz González1* 1 Departamento de Fisiología Vegetal, Universidad de Santiago de Compostela, 15782 Santiago de Compostela, Spain. Abstract The source of germplasm as well as the technique used for storage of mosses can enhance survival after cryopreservation. Samples of gametophores, protonemata and protonemal brood cells from in vitro cultures of Splachnum ampullaceum were cryopreserved following exposure to a plant vitrification solution (PVS2) for two different times (5 and 10 min) at 0şC. Half of the samples were pretreated with a loading solution containing 2 M glycerol and 0.4 M sucrose before exposure to PVS2. After one week storage in liquid nitrogen, S. ampullaceum samples were regenerated on Gamborg’s B5 mineral medium with B5 vitamins. Exposure to a loading solution was a prerequisite for high survival in all samples. Four weeks after cryopreservation, 92.3% brood cells, 60.0% gametophores and 46.0% protonemata pretreated with a loading solution had regenerated, displaying normal growth and development, thus demonstrating that vitrification is a useful method for moss cryopreservation. Keywords: bryophyte, moss, cryopreservation, vitrification, brood cells.
CryoLetters 31 (1), 29-39 (2010) CRITICAL MOISTURE CONTENT WINDOWS DIFFER FOR THE CRYOPRESERVATION OF POMELO (Citrus grandis) SEEDS AND EMBRYONIC AXES Bin Wen*, Chuantao Cai, Ruling Wang, Yunhong Tan and Qinying Lan Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences, Mengla, Yunnan, 666303, China. Abstract Cryopreservation was attempted using a partial dehydration and freezing protocol on pomelo (Citrus grandis) seeds of ten cultivars and embryonic axes of one cultivar collected from Xishuangbanna. Although seeds of all ten cultivars could be dehydrated safely to 10% moisture content, further dehydration impaired seed viability with critical moisture contents ranging from 9% to 7%. Complete seedling regenerated from seeds frozen in a moisture window between 5-9%, and maximum seedling recovery varied between 22-86%. Cryopreservation of Jiajieyou cv. embryonic axes had a moisture window between 3-20%, much wider compared to whole seed cryopreservation, and maximum post-thaw emergence of 93% was achieved at 13% moisture content. Emergence differed only slightly from survival when whole seeds were cryopreserved, but for the embryonic axis cryopreservation at low moisture contents resulted in much lower emergence than survival. Possible causes of intraspecific variation in cryotolerance in pomelo seeds is discussed. Keywords: Citrus grandis; cryotolerance; desiccation tolerance; intermediate seeds; intraspecific variation
CryoLetters 31 (1), 40-49 (2010) CRYOPRESERVATION OF FRASER PHOTINIA (Photinia x fraseri Dress.) VIA VITRIFICATION-BASED ONE-STEP FREEZING TECHNIQUES Yelda Ozden Tokatli* and Hülya Akdemir Gebze Institute of Technology, Faculty of Science, Department of Biology, 41400, Kocaeli, Turkey Abstract An efficient vitrification-based cryopreservation procedure was developed for Fraser photinia shoot apices by assessing the influences of various vitrification solutions (PVS1, PVS2, PVS3 and VSL) and different vitrification methods (cryovial vitrification, droplet vitrification and droplet freezing) on shoot regrowth. Moreover, influences of cold-hardening period (0 to 8 weeks), preculture medium (with sucrose and proline) and regrowth medium (QL plus 4.4, 8.8 and 17.6 µM BA) were also evaluated. Among the different procedures tested, best shoot regrowth (40.3%) was achieved by using a droplet vitrification technique in which cold-hardened and precultured shoot apices were vitrified for 120 min at 0°C in droplets, rapidly cooled, thawed and then cultured on 17.6 µM BA-containing QL medium. Overall results indicated the importance of not only the composition of vitrification solution, and preculture and regrowth media, but also the application of an appropriate vitrification technique to achieve optimum recovery post-cryopreservation. Keywords: cryovial vitrification, droplet vitrification, PVS1, PVS2, PVS3, VSL
CryoLetters 31 (1), 50-62 (2010) Effect of timing, DOSe and Interstitial Versus Nanoparticle delivery of tumor necrosis factor alpha in combinatorial adjuvant cryosurgery treatment of ELT-3 Uterine fibroid tumor Jing Jiang1 and John Bischof*1,2,3 Department of Biomedical Engineering1, Mechanical Engineering2 and Urologic Surgery3, University of Minnesota, Minneapolis, MN 55455, USA. Abstract Cryosurgery has shown potential as a minimally invasive technology for tumor treatment. However, incomplete destruction followed by tumor recurrence after cryosurgery is a common drawback. This study characterizes several variables in the cryoadjuvant TNF-α enhancement of conservative cryosurgery (i.e. freezing to the visible edge) of ELT-3 (uterine leiomyoma) tumor in a female nude mouse model. The variables include pretreatment time, mode of TNF-α delivery (native vs. CYT-6091, a PEGylated 33 nm colloidal gold core nanoparticle) and dose of TNF-α. Survival and tumor growth delay were measured up to 30 days and showed: 1) pretreatment with TNF-α required 4 hours incubation prior to cryosurgery to produce a tumor growth delay over cryosurgery alone, and 2) CYT-6091 reduced the toxicity of TNF-α administration over intratumoral or peritumoral injection of native TNF-α. Taken together, 5 µg TNF-α delivered by the nanodrug CYT-6091 4 hours prior to cryosurgery yielded a dramatic reduction in tumor growth over cryosurgery alone and in some cases even total remission of the tumor. However, some toxicity at higher doses (i.e. 5 µg) with CYT-6091 was noted compared to previous work in prostate (LNCaP) cancer grown in a male nude mouse. Potential reasons for this, including sex and weight of the animals are discussed. Further opportunities to optimize the TNF-α enhanced cryosurgical response of this tumor include dosing between 2 - 5 µg at 4 hours prior to cryosurgery, and freezing beyond the visible edge of the tumor. Keywords: Cryosurgery, TNF-α, CYT-6091, leiomyoma, drug delivery
CryoLetters 31 (1),63-75 (2010) CRYOPRESERVATION OF Fraxinus excelsior L. EMBRYOGENIC CALLUS BY ONE-STEP FREEZING AND SLOW COOLING TECHNIQUES E. A.Ozudogru1, M. Capuana2, E. Kaya1, B. Panis3 and M. Lambardi4* 1GYTE / Gebze Yuksek Teknoloji Enstitusu, Istanbul Caddesi, No 101, 41400, Gebze, Kocaeli, Turkey Abstract An efficient cryopreservation protocol for the safe storage of Fraxinus excelsior L. embryogenic callus cultures is reported. The cryopreservation methods tested included one-step freezing by means of (i) encapsulation-vitrification; or (ii) encapsulation-dehydration; and (iii) slow cooling using the Nalgene Freezing container, Mr Frosty®, which produces a temperature decrease of about 1şC min-1 when placed in a -70şC freezer. None of the one-step freezing techniques was effective for cryopreservation of encapsulated callus masses, irrespective of the cryoprotective treatment applied, i.e., treatment with the PVS2 vitrification solution or physical dehydration with silica gel before direct immersion in liquid nitrogen. On the contrary, when a slow cooling protocol was applied to embryogenic callus which had been pretreated for 60 min with a 210 g l-1 (0.61 M) sucrose-7.5% DMSO cryoprotective solution, up to about 1.3 g per Petri dish of proliferating callus was observed 42 days after recovery from liquid nitrogen, and cultures were able to produce somatic embryos 8 weeks after transfer to semi-solid medium. TTC staining of callus cultures provided a fast evaluation of culture viability. Keywords: common ash, encapsulation-dehydration, encapsulation-vitrification, slow cooling, somatic embryogenesis |
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Abstracts |
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For Abstracts published from meetings, such as SLTB meetings, go to the
relevant Volume Year of the journal (above). For Full text Free Access Content (from 2000 onwards) go to CryoLetters at Ingenta and look for the blue symbol. |
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