• Plant Biotechnology Reports
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• v.2 no.2
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• pp.123-131
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• 2008

Vitrification methods are convenient for cryopreserving plant specimens, as the specimens are plunged directly into liquid nitrogen (LN) from ambient temperatures. However, tissues and species with poor survival are still not uncommon. The development of vitrification solutions with high survival that cover a range of materials is important. We attempted to develop new vitrification solutions using bromegrass cells and found that VSL, comprising 20% (w/v) glycerol, 30% (w/v) ethylene glycol, 5% (w/v) sucrose, 10% (w/v) DMSO and 10 mM $CaCl_2$, gave the highest survival following cryopreservation, as determined by fluorescein diacetate staining. However, the cryopreserved cells showed little regrowth, for unknown reasons. To check its applicability, VSL was used to cryopreserve gentian axillary buds and the performance was compared with those of conventional vitrification solutions. Excised gentian stem segments with axillary buds (shoot apices) were two-step precultured with sucrose to induce osmotic tolerance prior to cryopreservation. Gentian axillary buds cryopreserved using VSL following the appropriate preculturing approach exhibited 78% survival (determined by the regrowth capacity), which was comparable to PVS2 and PVS1 and far better than PVS3. VSL had a wider optimal incubation time (20-45 min) than PVS2 and was more suitable for cryopreserving gentian buds. The optimal duration of the first step of the preculture was 7-11 days, and preculturing with sucrose and glucose gave a much higher survival than fructose and maltose. VSL was able to vitrify during cooling to LN temperatures, as glass transition and devitrification points were detected in the warming profiles from differential scanning calorimetry. VSL and its derivative, VSL+, seem to have the potential to be good alternatives to PVS2 for the cryopreservation of some materials, as exemplified by gentian buds.

• Proceedings of the Korean Radioactive Waste Society Conference
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• 2003.11a
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• pp.524-531
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• 2003

The usefulness of vitrification technology for low- and intermediate- level radioactive wastes was demonstrated because of high volume reduction, mechanical and chemical stability of final waste forms. Thus, a construction of the commercial vitrification plant Is currently promoted. Due to the high radiation level of the waste, shielding analysis has to be carried out for safe design in a vitrification facility. Because there has been no experience in the construction and operation of the vitrification facility in Korea, in this study, in order to get some information for help the detailed design and plan for operation in vitrification facility, shielding analysis for each facility in pilot plant is carried out by using source term from established study. For the selection of the shielding material, concrete was chosen compared to the lead because of economic advantage, weight consideration, and thermal resistance.

• Journal of Nuclear Fuel Cycle and Waste Technology(JNFCWT)
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• v.8 no.2
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• pp.143-150
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• 2010

Vitrification technology has been gradually recognized as one of effective solidification methods for concentrated boric acid wastes generated in PWR. Vitrification for low- and intermediate-level radioactive wastes has a large volume reduction and good durability for the final products. A feasibility study for the vitrification of concentrated boric acid wastes has been performed with developing the pre-treatment methods of powdered wastes, glass compositions using glass formulation and demonstration test. The pre-treatment method is pelletizing the powder type for stable feeding within cold crucible melter. The glass compositions should be developed considering molten glass are related with wastes reduction. High contents of sodium and boron within borate wastes give influence to waste loading. A variety of factors obtained from the demonstration test are reviewed, which is wastes feeding rate, off-gas characteristics on stack and glass characteristics of final products such as durability for implementing the wastes disposal requirement. The aim of this paper is to present the feasibility of vitrification and review the solidification method for concentrated boric acid wastes and obtain the physicochemical characteristics of solidified glass.

• Korean Journal of Veterinary Research
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• v.35 no.3
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• pp.635-641
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• 1995

The purpose of this study was to investigate the effects of developmental stage and equilibration time on survival of rabbit embryos following freezing by vitrification. Adult New Zealand White female rabbits were superovulated with PMSG and hCG. The 8-cell stage embryos were collected from 40 to 45 hours after hCG injection by flushing oviducts with Dulbecco's phosphated buffered saline and in vitro cultured in TCM-199 containing 10% fetal calf serum(FCS). Each embryos developed in vitro to 16-cell, compact morula and blastocyst was cryopreserved and cultured following thawing to examine their developmental potential to expanded blastocyst stage in vitro. The frozen-thawed-cultured embryos were stained with Hoechst 33342, and their nuclei were counted using a fluorescence microscope. On the toxicity test of EFS solution as cryopreservation, the survival rates of 8-cell stage embryos was decreased in reverse to increasing of exposure time over 5 minutes. The post-thaw survival rates of embryos on equilibration times was significantly(P<0.05) higher for 2 min. than for 5 or 10 minutes. From morula to blastocyst of rabbit embryos was more suitable than 8-cell stage for cryopreservation by vitrification. The higher post-thaw survival rate of embryos can be achieved by keeping the cryoprotectant at $4^{\circ}C$ than at $20^{\circ}C$. The mean number of nuclei per embryo following freezing by vitrification and in vitro culture to expanded blastocyst at compacted morula and blastcyst was not significantly differ from fresh blastocyst.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2021.04a
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• pp.36-36
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• 2021

Cryopreservation has been broadly used as an efficient method for a long-term conservation for many types of plants especially vegetatively propagated plants. Among several cryopreservation methods, a droplet-vitrification was the most widely applicable and efficient method. Studies have developed protocols for strawberry using droplet-vitrification method and suggested the practical use of the protocol for large number of germplasm with a little modification. In this study, the droplet vitrification method of shoot tip has been tested on 31 accessions provided around the world. Shoot tips were precultured on Murashige and Skoog (MS) liquid medium supplemented with 0.3~0.5M sucrose. Precultured explants were osmoprotected with loading solution, 35% of PVS3 (C4, 17.5% glycerol and 17.5% sucrose) for 40 min and exposed to dehydration solution, PVS3 (B1, 50% glycerol and 50% sucrose) for 60 min. Then, the explants were transferred onto droplets containing 2.5 uL PVS3 on sterilized aluminum foils prior to direct immersion in liquid nitrogen (LN) for 1hr. The cryopreserved shoot tips were rapidly warmed in a water bath at 40C and then unloaded in MS with 0.8M sucrose for 40 min. The shoot tips were cultured in NH₄NO₃-free MS post culture medium for 2 weeks. Subsequently, the explants were moved to the MS medium for 6 weeks and evaluated the regrowth rate. By this droplet-vitrification protocol, twenty-four accessions showed at least 40% regrowth rate. Out of 24 accessions, 'Nonsan1ho' had the highest regeneration rate of 85.8% and 'Jumbo pureberry' had the lowest with 42.1%.

• Korean Journal of Plant Resources
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• v.36 no.6
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• pp.571-579
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• 2023

In this study, cryopreservation by droplet-vitrification was applied to pear (Pyrus spp.) germplasm. We focused on the development and assessment of various strategies for the selection of suitable tissue, osmoprotection, and dehydration. We also evaluated post-thaw recovery of cryopreserved explants by droplet-vitrification. Preferentially, we tested the effects of preculture and loading treatments to determine which tissues were more suitable, either the apical shoot tips or the axillary buds. Apical shoot tips showed the better regrowth rate than in vitro axillary buds. The most effective techniques for cryopreservation were as follows. Shoots from in vitro seedlings which had been cultured for about 5-6 weeks were cold-hardened at 4℃ for one week, excised shoot tips were precultured on liquid MS medium including 0.3 M sucrose for 31 hours and 0.7 M sucrose for 17 hours, osmoprotected in loading solution (LS) for 40 min, and then cryoprotected in dehydration solution (PVS3) for 90 min. In addition, we found that regrowth rates of explants on regrowth medium after exposure to liquid nitrogen (LN) were higher than those on MS medium. Results indicated that the highest regrowth percentage was 95.6% for 'Bartlett' cultivar and 68.9% for 'BaeYun No.3' cultivar. Consequently, apical shoot tips of two pear cultivars, 'Bartlett' (P. communis) and 'BaeYun No.3' (P. pyrifolia), were successfully cryopreserved by droplet-vitrification. Results of this study show that the enhanced droplet-vitrification method described in the present study could be used as an effective means for long-term storage of pear genetic resources.

• Journal of Embryo Transfer
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• v.20 no.2
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• pp.129-135
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• 2005

The present study examines the effects of kinds and concentrations of cryoprotectants, PVP and sucrose and trehalose on the survival rate of vitrified porcine embryos. 1. The developmental stages for the embryos used in vitrification were $245(23.0\%)$ for 2 cell stage, $256(24.1\%)$ for the blastocyst, $234(22.0\%)$ for the early blastocyst $221(20.8\%)$ from the expanded blastocyst and $107(10.1\%)$ from hatching blastocyst out of 1,063 embryos. 2 The survival rate of morula, early blastocyst and expanded blastocyst vitrification-thawed with EDT and EGS were $69.1\%,\;70.3\%,\;69.8\%\;and\;62.5\%,\;61.7\%,\;63.6\%$, respectively. The expanded blastocyst treated with EDS showed the highest survival rate compared with the other cryoprotectants. 3. The survival rate of early blastocyst, expanded blastocyst and hatching blastocyst vitrification-thawed with EDS diluted in $medium + 10\%$ FCS were $61.1\%,\;27.8\%,\;16.7\%$, respectively. This result were love. than the control of group $(92.3\%,\;71.2\%,\; 55.8\%)$. 4. The survival rate of embryos vitrified with EDS and EDT supplemented with $10\%\;and\;20\%$ PVP were $74.3\%,\;77.5\%\;and\;79.4\%,\;71.1\%$, respectively. The survival rate of vitrified embryos cultured for $24\~48$ hours were $37.1\%,\; 40.0\%\;and\;35.3\%,\;31.6\%$ which were significantly lower than that of non-cultured embryos. The survival rate of embryos vitrified with EDS and EDT supplemented between $10\%\;or\;20\%$ PVP did not have a significant difference. 5. The survival rate of embryos vitrification-thawed with EDS to morula, early blastocyst, expanded blastocyst and hatching blastocyst were $58.2\%,\; 36.4\%,\;14.5\%$ to morula, $62.5\%,\;45.8\%,\;20.8\%$ to early blastocyst, $74.1\%,\;61.1\%,\;29.6\%$ to expanded blastocyst and $60.0\%,\;40.0\%,\;14.0\%$ to hatching blastocyst.

• Korean Journal of Animal Reproduction
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• v.23 no.4
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• pp.313-321
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• 1999

The purpose of these study was to investigate the use of a glass micropipette (GMP) as a vessel for vitrification of bovine IVP blastocysts, to compare the post-thaw survival rates of bovine blastocysts frozen in GMP with those frozen in OPS that have been previously investigated, and to improve the hatching rate following vitrification with GMP method. The GMP vessel permits higher freezing and warming rate than the OPS due to the higher heat conductivity of the glass and lower mass of the solution that contains the embryos. Groups of three bovine IVP blastocysts were sequentially placed into vitrification solution before being loaded into either the OPS or GMP vessels and immersed into L$N_2$within 20 to 25 sec. Post-thaw blastocysts were serially washed in 0.25 and 0.15 M sucrose in HM and TCM-199 for each 5 min, respectively, and then cultured in TCM 199 supplemented with 10% FCS for 24 h. The rate of blastocyst re-expanding did not significantly different for OPS (75.9%) and GMP (80.0%) methods (P＞0.05). The hatching rates in OPS (34.1%) and GMP (37.5%) methods were significantly lower than that in control group (54.3%) (P＞0.05). In addition, the rate of blastocyst re-expanding was significantly lower if blastocysts were vitrified in the wide portion of the micropipette rather than the narrow portion of the micropipette (83.3 vs 56.7%) (P＞0.05), even though three blastocysts were loaded per vessel. The hatching rate in 0.05% pronase solution treatment for 30, 60 and 90 see (45.9, 54.7 and 57.5%) were significantly higher than that in control (35.0%), even though there was not significantly different between 30 see and control. These results indicate that both vitrification vessels can provide high survival rates of bovine IVP blastocysts. However, the GMP vessel has the advantage over the OPS, in that the former does not need a cap to protect the vessel from floating after immersion in L$N_2$. The location of the embryos (narrow or wide portion of immersion) were considered to be limiting factors to the viability of bovine IVP embryos. The exposing in 0.05% pronase solution for 60 or 90 see can increase hatching rates of post-thaw bovine IVP blastocysts.

• Proceedings of the Korean Society of Crop Science Conference
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• 2005.04a
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• pp.304-305
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• 2005

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2003.10a
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• pp.113-113
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• 2003