• Fisheries and Aquatic Sciences
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• v.27 no.5
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• pp.306-313
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• 2024

The genus Chirostoma is endemic from the Mesa Central of Mexico. It is conformed of 18 species and six subspecies. Five species are in some category of risk, because of this, Chirostoma jordani is an excellent model species to implement biotechnologies like gametes cryopreservation. Aim of present study was to evaluate fertilizing capacity of cryopreserved C. jordani sperm, as alternative to conservation and assisted reproduction in this specie and genus. Males and females were collected from wild Atlangatepec dam stock, Tlaxcala State, Mexico. Seminal quality was evaluated in fresh and cryopreserved semen with three cryoprotective agents (CPAs): 10% dimethyl sulfoxide (DMSO), 10% methanol (MeOH), 14% ethylene glycol (EG) and it was determined its post-thaw fertilizing capacity. Sperm motility percentage decreased during cryopreservation process (p < 0.05). There were not significant differences in post-thaw motility percentage between EG (53.5 ± 1.9%) and MeOH (53.3 ± 1.3%), but DMSO (50.3 ± 0.5%) was significantly different (p < 0.05). Results showed that 0.2 μL fresh semen were enough to fertilize 100% oocytes (n = 60). 10 μL DMSO and 5 μL MeOH and EG cryopreserved semen were necessary to fertilize oocytes 100% (n = 60) (p < 0.05). Cryopreservation and fertilization protocol for C. jordani sperm was efficient and it could be used for its assisted reproduction.

• Clinical and Experimental Reproductive Medicine
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• v.37 no.4
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• pp.307-319
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• 2010

Objective: Vitrification requires a high concentration of cyroprotectant (CPA) and an elevated cooling speed to avoid ice crystal formation. We have evaluated the effect of different combinations of cooling rate and CPA on embryonic integrity (developmental competence) in order to increase the efficiency of vitrification without impairing embryo viabilit. We hypothesized that the combination of CPA or the increase of cooling rates can reduce the concentration of toxic CPA for vitrification. As consequently, we performed experiments to evaluate the effect of various composition of CPA or slush nitrogen ($SN_2$) on the mouse embryonic development following vitrification using low CPA concentration. Methods: Vitrification of mouse embryos was performed with EM grid using liquid nitrogen ($LN_2$) or $SN_2$ and different composition of CPAs, ethylene glycol (EG) and dimethylsulfoxide (DMSO). After vitrification-warming process, their survival and blastocyst formation rates were examined. For analyzing long-term effect, these blastocysts were transferred into the uterus of foster mothers. Results: Survival and blastocyst formation rates of vitrified embryos were higher in EG+DMSO group than those in EG only. Furthermor, the group using $SN_2$ with a lower CPA concentration showed a higher survival of embryos and developmental rates than group using $LN_2$. Conclusion: The combination of EG and DMSO as CPAs may enhance the survival of mouse embryos and further embryonic development after vitrification. $SN_2$ can generate high survival and developmental rate of vitrified/warmed mouse embryos when a lower concentration of CPA was applied. Therefore, these systems may contribute in the improvement of cryopreservation for fertility preservation.

• Journal of Microbiology and Biotechnology
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• v.28 no.5
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• pp.718-731
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• 2018

The beneficial effects of lactic acid bacteria (LAB) have been intensively investigated in recent decades with special focus on modulation of the host intestinal microbiota. Numerous discoveries of effective probiotics are driven by a significantly increasing demand for dietary supplements. Consequently, technological advances in the large-scale production and lyophilization are needed by probiotic-related industries for producing probiotic LAB for commercial use. Our study had a dual objective, to determine the optimum growth medium composition and to investigate appropriate cryoprotective additives (CPAs) for Lactobacillus salivarius, and compare its responses with other Lactobacillus species. The one-factor-at-a-time method and central composite design were applied to determine the optimal medium composition for L. salivarius cultivation. The following composition of the medium was established (per liter): 21.64 g maltose, 85 g yeast extract, 1.21 ml Tween 80, 6 g sodium acetate, $0.2g\;MgSO_4{\cdot}7H_2O$, $0.02g\;MnSO_4{\cdot}H_2O$, $1g\;K_2HPO_4$, $1.5g\;KH_2PO_4$, $0.01g\;FeSO_4{\cdot}7H_2O$, and 1 g sodium citrate. A cryoprotective additive combination comprising 10% (w/v) skim milk and 10% (w/v) sucrose supplemented with 2.5% (w/v) sodium glutamate was selected for L. salivarius, and its effectiveness was confirmed using culture-independent methods in the freeze-dried cells of the Lactobacillus strains. In conclusion, the optimized medium enhanced the species-specific cultivation of L. salivarius. On the other hand, the cryoprotective effects of the selected CPA mixture may also be dependent on the bacterial strain. This study highlights the necessity for precise and advanced processing techniques for large-scale production of probiotics in the food and feed industries.

• The Korean Journal of Malacology
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• v.29 no.3
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• pp.251-258
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• 2013

This study aims to find out a suitable cryoprotective agent (CPA) for cryopreservation and its optimum concentration in order to conduct planned artificial seed production of Pacific oyster, Crassostrea gigas and to preserve superior sperm. For this, we tried to understand toxicity and the effect of cryopreservation by CPA type and concentrations first and then looked into cell damage of the sperm after thawing. Toxicity analysis on the sperm of Pacific oyster according to different CPA and immersion time shows that dimethyl sulfoxide (DMSO) comes first when it comes to survival rate and mobility followed by ethylene glycol (EG), glycerol and methanol. To identify the optimum CPA and its level, filtered seawater was used as a diluent before cryopreservation for 30 days. As a result, cryopreserved sperm of Pacific oyster with 15% of DMSO showed the highest survival rate and activation. Also, we observed the cryopreserved and thawed sperm with Scanning electron micrographs by CPAs and concentrations. Consequently, DSMO showed the lowest cell damage followed by EG, methanol, glycerol and the level was 15, 20, 10, 5% respectively. In a nutshell, it is proven that the optimum CPA and its level is 15% of DMSO.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2019.04a
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• pp.62-62
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• 2019

Bacterial leaf blight(BLB), caused by X. oryzae pv. oryzae(Xoo), is one of the most destructive diseases of rice due to its high epidemic potential. Understanding BLB resistance at a genetic level is important to further improve the rice breeding that provides one of the best approaches to control BLB disease. In the present investigation, a collection of 192 accessions was used in the genome-wide association study (GWAS) for BLB resistance loci against four Korean races of Xoo that were represented by the prevailing BLB isolates under Xoo differential system. A total of 192 accessions of rice germplasm were selected on the basis of the bioassay using four isolated races of Xoo such as K1 and K2. The selected accessions was used to prepare 384-plex genotyping by sequencing (GBS) libraries and Illumina HiSeq 2000 pairedend read was used for GBS sequencing. GWAS was conducted using TASSEL 5.0. The TASSEL program uses a mixed linear model (MLM). The results of the bioassay using a selected set of 192 accessions showed that a large number of accessions (93.75%) were resistant to K1 race and K2 resistant germplasm proportion remained between 66.67. The genotypic data produced SNP matrix for a total of 293,379 SNPs. After imputation the missing data was removed, which exhibited 34,724 SNPs for association analysis. GWAS results showed strong signals of association at a threshold of [-log10(P-value)] more than 5 (K1 and K2) for nine of the 39 SNPs, which are plausible candidate loci of resistance genes. These SNP loci were positioned on rice chromosome 2, 9, and 11 for K1 and K2 races. The significant loci detected have also been illustrated and make the CPAS markers for NBS-LRR type disease resistance protein, SNARE domain containing protein, Histone deacetylase 19, NADP-dependent oxidoreductase, and other expressed and unknown proteins. Our results provide a better understanding of the distribution of genetic variation of BLB resistance to Korean pathogen races and breeding of resistant rice.

• Journal of Marine Life Science
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• v.8 no.2
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• pp.115-120
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• 2023

This study aims to find out a suitable extender and cryoprotective agent (CPA) for cryopreservation and its optimum concentration in order to conduct planned artificial seed production of Korean bullhead Pseudobagrus fulvidraco and to preserve superior sperm. Experiments were designed to investigate the effects of the different combinations of three extenders (I: 300 mM glycose, II: Kurokura extender, III: Li extender), four cryoprotectants (dimethyl sulfoxide, ethylene glycol, methanol and glycerol) and four concentrations (5, 10, 15, 20%) on the cryopreservation of Korean bullhead sperm. Postthawed sperm survival rate and sperm activity index (SAI) were detected to evaluate the effects of sperm cryopreservation. The optimal combination of extender and CPA for cryopreservation of Korean bullhead sperm was extender III + 10 and 15% methanol, resulting in a survival rate and SAI of 66.9 ± 8.7, 67.3 ± 13.1% and 2.6 ± 0.4, 2.6 ± 0.5 respectively, which was higher than had been achieved with other extenders and CPAs.