Volcanic seawater has been naturally filtered and purified by volcanic rock layers. It contains abundant minerals such as calcium, magnesium, and iron. This study investigated the genotoxicity of calcium from Jeju lava seawater (CJLS). We performed bacterial reverse mutation assay, chromosomal aberration assay, and mammalian micronucleus test at up to 5,000 g/plate concentrations with or without metabolic activation to determine the CJLS genetic toxicity. None of these tests showed any mutagenic potential. The bacterial reverse mutation assay showed that the CJLS did not induce mutagenicity in Salmonella typhimurium TA98, TA100, TA1535, TA1537, and Escherichia coli WP2uvrA with or without metabolic activation of the S9 mixture. The oral administration of CJLS also did not significantly increase the number of micronucleated polychromatic erythrocytes or the mean ratio of polychromatic to total erythrocytes. Additionally, CJLS did not cause a significant chromosomal aberration in CHL cells in the presence or absence of S9 activation. Therefore, CJLS could be considered as a reliable and safe functional food ingredient.

Upgraded activated carbons (ACs) are typically synthesized by mixed methods, such as solid–solid mixing and wet impregnation of low-grade ACs with KOH. This study compares the properties of upgraded ACs prepared by different methods using elemental analysis, X-ray photoelectron spectroscopy, N2 adsorption isotherms, and X-ray diffraction. In ACs produced by the solid–solid mixing, the ratio of potassium activator is proportional to the surface area and amount of gas produced. However, in wet impregnated ACs, the potassium ratio exhibits a zero or negative correlation. It is demonstrated that potassium ions in solution are not transferred to K2O and do not contribute to the surface area and pore size, generating less amount and different composition of gases. As such, impregnated ACs exhibit similar surface areas and large pores, regardless of the potassium ratio. The physical properties, such as specific surface areas and pore size distribution, of ACs using wet impregnation were similar to the ACs generated by the water physical activation. It indicated that the KOH does not efficiently act as a chemical activator in the wet impregnation method. Therefore, a certain amount and suitable mixing method of chemical activator play an important role in the property upgrade of ACs.

To prepare activated carbon with a high specific surface area, oxygen functional groups (OFGs) that can serve as useful electron donors during KOH activation were treated with nitric acid and incorporated into activated carbon. OFGs are incorporated differently according to the surface characteristics of starting materials. Up to 22.46% OFGs are incorporated into wood-based activated carbons (WACs), the C=O, COOH contents was 1.90, 17.05%, respectively. Whereas up to 12.82% OFGs are incorporated into coconut shell-based activated carbons, the C=O, COOH contents was 4.12, 6.15%, respectively. The OFGs used for increasing the specific surface area are the carbonyl group, and as the content of the functional group increases, the carbonyl group spreads to the carboxyl group. The specific surface area of activated carbons increased by 10–68% with an increase in the carbonyl group up to 6% (maximum point of carbonyl group). On the other hand, the specific surface area for WACs increased when the carboxyl group was 10% or below, but decreased by 6–15% when it increased to 10% or excess.

In this study, commercial activated carbons (ACs) were upgraded by different activation methods, and the gases generated during the activations were defined and quantified. The chemical activation commonly applied for upgrading ACs uses complex reactions, involving pyrolysis, physical, and chemical reactions. The ACs based on wood materials were characterized by elemental analysis, N2 physisorption, Fourier-transform infrared spectroscopy, X-ray photoelectron spectroscopy, and temperature-programmed desorption mass spectrometry. The patterns and composition of the generated gases were analyzed by gas chromatography and X-ray diffraction; high-resolution scanning electron microscopy was also used to characterize the activated carbon. The AC was mostly decomposed to CO2 by pyrolysis and physical activation, while CO was mainly detected during chemical activation from the K2CO3 produced by the reactions between CO2 and K2O. The detected amounts of generated gases were differed at various KOH ratios and residence times. The highest surface area obtained in this study was 2000 m2/g at the optimum ratio of AC and KOH (1:2).

In the present study, we investigated the effect of porcine follicular fluid (PFF) concentration (10% vs. 1%) and protein-free media (PFF 0%) on maturation of porcine oocytes in vitro and analysed difference in gene expression in resulting blastocysts following parthenogenetic activation. Three groups were tested; 1) 10% PFF: Tissue culture medium (TCM) 199+10% PFF; 2) 1% PFF: TCM 199+1% PFF; and 3) 0.1% PVA: TCM 199+0.1 PVA. Cumulus-oocyte-complexes were cultured in the respective media containing gonadotrophin (1 ug/ml), epidermal growth factor (10 ng/ml), cystein (0.57 mM), sodium pyruvate (0.91 mM), insulin (5 ug/ml), 9-cis retinoic acid (5 nM) for 20～22 h and then without hormonal supplements for an additional 20-22 h. Data was analyzed using statistical analysis system(SAS) program. There was no significant difference in oocyte maturation rate. However, significantly higher (p<0.05) proportions of embryos developed to the blastocyst stage when oocytes were matured in 10% PFF group (45%) than in the 1% PFF group (31.1%). The total cell numbers were not significantly different among groups (52 ± 1.3 vs. 54.6±3.1 vs. 54.4±2.5, respectively). The relative abundance (ratio to beta-actin mRNA) of gene transcripts related to apoptosis in blastocysts was measured by real- time PCR. The expression of anti-apoptotic gene (BclxL) was up-regulated and the expression of pro-apoptotic gene (Bax) was down-regulated in 10% PFF group than in the other groups. Therefore, it can be concluded that supplementation of 10% PFF during in vitro maturation improves embryo development by reduction of apoptosis. * This study was supported by IPET (#311011-05-1-SB010), RNL Bio (#550-20120006), MKE (#10033839-2011-13), Institute for Veterinary Science, the BK21 program and TS Corporation.

Embryo transfer (ET) is the final procedure for getting pregnancy through assisted reproductive technology such as IVF (in vitro fertilization), SCNT (somatic cell nuclear transfer). In our laboratory, the porcine cloned embryos loaded in ET medium are carried for 3 hours by portable incubator because of the great distance from the laboratory to the experimental farm. Thus, before transferring into recipient, porcine cloned embryos are exposed in vitro condition for long time. Medium which is used in this process is the TALP (Tyrode’s medium supplemented with 10 mM HEPES), but it includes little nutrients for embryo. Thus, the aim of this study is to determine whether ET media containing nutrients affect the in vitro development of embryos compared to TALP. For the experiment, porcine zygote medium (PZM)-5 which has amino acids for developing embryo was chosen as ET medium containing nutrients, added 10 mM Hepes as PZM-5 does not contain buffering system. For experiment, we carried out parthenogenesis through a chemical method using Thi/DTT. Parthenogenetic embryos were cultured in PZM-5 for 2 days, and then they were randomly divided into two group; loaded in a straw with TALP or PZM-5-Hepes, respectively. They were stored in a portable incubator for 3 hours to simulate the time consumed in ET, thereafter embryos in both TALP and PZM-5-Hepes groups were respectively cultured in PZM-5 for additional 5 days. All experiments were repeated 5 times. In result, blastocyst formation rate were 22.46%±1.47 and 23.17%± 2.13, respectively and total cell number were 32.9±2.22 and 37.09±2.18, respectively. There is no significant difference between TALP and PZM-5-Hepes groups. * Further study will investigate effect of PZM-5-Hepes on in vivo development of porcine cloned embryo. This study was supported by IPET (#311011-05-1-SB010), RNL Bio (#550-20120006), Institute for Veterinary Science, the BK21 program and TS Corporation.

Adipose tissue-derived mesenchymal stem cells (ASCs) are very interesting in several laboratory animals and humans because they are easy to harvest and expand to generate millions of cells from a small quantity of fat. ASCs are known as useful materials for clinical applications in human cell therapy and as a donor cell in somatic cell nuclear transfer (SCNT). Here, we investigated if 1) minipig ASCs can be isolated, self-renewed and differentiated into multiple tissue lineages, 2) ASCs can be a suitable donor cell type for generation of cloned pig. In order to isolate ASC, adipose tissues were collected from inguinal region of a 6-year-old female minipig. The ASCs were attached to the culture dish with a fibroblast-like morphology. They expressed cell-surface marker characteristics of stem cell, underwent osteogenic, adipogenic, myogenic, neurogenic and chondrogenic differentiation when exposed to specific differentiation-inducing conditions. To investigate its potential as donor cell for cloning, we respectively carried out SCNT using ASC, adult skin fibroblast (ASF) and fetal fibroblast (FF) derived from same minipig. The ratio of blastocysts to 2-cell embryos and total cell number of blastocysts were monitored as experimental parameters. In results, cleavage and developmental competence to blastocysts rate showed no significant difference among the three groups. On the other hand, total cell numbers of blastocysts derived from ASC and FF were significantly higher than in ASF (89±7.9 and 105±5.5 vs. 57.5±5.2, respectively). Our results demonstrated that ASC have potential compared to ASF and FF in terms of the in vitro development and blastocyst formation ability. In further study, we will investigate the in vivo developmental ability of ASC as donor cell for pig cloning. * This study was supported by IPET (#311011-05-1-SB010), RNL Bio (#550-20120006), Institute for Veterinary Science, the BK21 program, TS Corporation and Optifarm Solution.

쌀국수 제품의 품질을 향상시키기 위한 방법으로서 기존의 쌀국수에 파스타가 지닌 물성 및 조리 상의 장점을 접목하고자 파스타의 제조원료인 세몰리나를 글루텐이 소량 첨가된 쌀가루에 대해 함량을 달리하여(0-20%, w/w) 대체하여 생면을 제조하였다. 제조된 생면을 끓인 조리면의 품질을 측정한 결과 전분노화가 다소 억제되며, 씹는 맛(“aldente”)이 증가하고, 끓이는 과정에서 일어나는 전분유출이 감소하여 전반적으로 제품의 품질이 향상되었다. 생면의 제조과정에서는 반죽의 점성 이 커져 반죽이 잘 뭉치고 단단해져 면대형성이 향상되는 효과가 기대되었다. RVA 측정에 따른 전분혼합액의 호화개시온도(pasting time)는 10% 첨가까지는 거의 영향이 없다가 20% 첨가구에서 최고 1.2 증가한 반면 최고점도(peak viscosity)는 첨가량 15% 이상부터 큰 폭으로 감소하였다. 전분의 breakdown값 역시 세몰리나 첨가량이 늘수록 급격히 줄어 shear thinning이 줄어들며 10% 이상 첨가구에서 최종점도(final viscosity)와 setback값 차이가 크게 줄어 노화억제에 의해 조리면이 딱딱해지는 정도가 감소하는 효과가 예상되었다. 생면을 끓일 때 국수에 의한 수분흡수가 낮아져 조리면의 중량 및 부피 변화가 감소되었으며 국물의 탁도가 증가되지 않아 전분침출 억제기능이 확인되었다. 조리면의 색도에서 명도(L)는 감소하였으며 적색도(a)와 황색도(b)가 증가하였으나 일정 수준(15%) 이상이 되면 그 영향이 적게 나타났다. 적은 양(5%)의 세몰리나 첨가에 의해서도 Texture Analyzer로 측정된 경도(hardness), 탄력성(springiness), 응집성(cohesiveness), 검성(gumminess) 및 씹힘성(chewiness)과 같은 모든 텍스쳐 특성치 및 인장강도가 소량(5%)의 첨가에 의해서도 현저히 증가하였으나 10% 이상에서는 변화폭이 줄어는 공통적인 변화양상을 보였다. 이상의 결과를 토대로 세몰리나를 첨가한 쌀국수를 제조하는 경우 원료비용 및 기능성을 고려할 때 쌀가루:세몰리나 최적 첨가비율은 9:1(10%, w/w)로 제시되었다.

The present study investigated the effects of follicle stimulating hormone (FSH) and human chorionic gonadotrophin (hCG) on the nuclear maturation of canine oocytes. Oocytes were recovered from mongrel female ovaries in various reproductive states; follicular, luteal or anestrous stage. Oocytes were cultured in serum-free tissue culture medium (TCM)-199 supplemented with various concentrations of FSH (Exp. 1: 0, 0.5, 1.0 or 10 IU) or hCG (Exp. 2: 0, 0.5, 1.0 or 10 IU) or both (Exp. 3: 1 IU FSH + 1 IU hCG) for 72 hr to determine the effective concentration of these hormones, and to examine their combined effect. After maturation culture, oocytes were denuded in PBS containing 0.1% (w/v) hyaluronidase by gentle pipetting. The denuded oocytes were stained with 1.9 μM. Hoechst 33342 in glycerol and the nuclear state of oocytes was evaluated under UV light. More (p<0.05) oocytes matured to MII stage when follicular stage oocytes were supplemented with 1 IU FSH (6.2%) compared with the control, 0.1 or 10.0 IU FSH (0 to 1.2%). Significantly higher (p<0.05) maturation rate to MII stage was observed in follicular stage oocytes supplemented with 1.0 IU hCG (7.2%) compared with the control or other hCG supplemented groups (0 to 1.5%). However, the combination of FSH and hCG did not improve the nuclear maturation rate of canine oocyte (2.4 %) compared with FSH (6.2%) and hCG alone (7.2%). In conclusion, FSH or hCG alone significantly increased the maturation of canine oocytes to MII stage.

온실 효과로 인한 지구온난화 현상은 전세계적으로 문제가 되고 있고, 온실효과를 일으키는 주원인은 온실가스이다. 온실 가스는 에너지 분야에서 가장 많이 배출되며, 2014년 기준 배출량의 86.8%를 차지한다. 배출 국가 온실가스 감축 의무부담에 관한 대응을 위해 에너지 사용 절감에 대한 전략이 동반되어 짐으로서, 에너지 절감 기술 및 소재 개발이 필요시 되었다. 국내 에너지 다소비 산업의 하나인 전해제련 공정 또한 여기서 벗어나지 못한다. 전해제련 공정은 수용액 전해조에 전극을 담그고 일정한 전류 혹은 전압을 가하여 수용액 속의 이온을 금속으로 석출하는 공정으로, 제조경비 중 전력비 비중이 높은 대표적인 에너지 다소비산업이다. 대표적 전해제련 생산품인 아연(Zn)은 최근 국제가격 하향안정화 추세로 국내기업의 글로벌 시장경쟁력 악화가 예상되며 이에 따른 가격 경쟁력 확보 필요성이 증가하였다. 본 연구에서는 Ir-Ta-Sn-Pd/Ti 전극을 이용해 아연 전해제련 시 사용되는 Pb 전극과 비교 하였고, 전류 밀도 500A/m2 조건에서 전위차 변화를 통해 전력소비 감소량을 예측하였다. 또한 아연 회수량 및 전극 표면 부식성 또한 관찰하여 Pb 전극 대체 효과를 확인하였다.

Spatiotemporal expressions of microRNAs (miRNAs) are altered by the physiological states of cells which could be influenced by microenvironment. Function of miRNAs has been focused as a new regulator of gene expressions and cell differentiation in human health and diseases. We found and identified the several miRNAs, which were related to developmental competence of preimplantation and implantation process of mouse blastocysts and outgrowth embryos by microarray-based bioinformatical studies. In this study, we evaluated three miRNAs expressions related to third cleavage event in conditioned media (CM) and blastocysts. Mouse 2-cell stage embryos were collected and monitored for 9 hours. The embryos were divided two groups as early third cleavage before 9 hours of collection and late third cleavage after 9 hours of collection. They were cultured to blastocyst stage up to day-5 after hCG injection. The total number of cells and the number of cells with fragmented DNA were assessed in blastocysts by terminal dUTP nick-end labelling (TUNEL) staining and DAPI staining. Mean cell number of early third cleavage group was significantly higher than that of late third cleavage group (105.3±8.0 vs 81.8±7.0, p<0.05), but apoptotic index was not different. The miRNAs of CM and blastocysts from early and late group were prepared, and quantified by qRT-PCR with TaqMan probes. The expression levels of three miRNAs (mmu-let-7b, mmu-miR-183, and mmu-miR-429) in CM and blastocysts were slightly upregulated in late third cleavage group. Our study suggested that the expression level of miRNAs could be altered with embryo quality, and miRNAs in CM may be used to predict miRNAs expression of embryos and developmental competence.

Discovery, identification, and informatics of low molecular weight peptide are extensively rising in the field of proteomics research. In this study, we analyzed protein profiles to discover peptide based biomarker for twelve different soybean seeds with three different agronomic types using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS). For optimization of SELDI-TOF MS in soybean seed proteome analysis, four different extraction buffers were tested with urea solubilization buffer, thiourea/urea solubilization buffer, phenol extraction buffer, and modified trichloroacetic acid (TCA)/acetone precipitation/urea solubilization extraction buffer. Two different type of ProteinChip arrays, cation exchange (CM10) and anion exchange (Q10), applied to profile peptides. Among the four different extraction buffers, phenol extraction was selected to protein extraction methodology. Numbers of detected peak cluster in twelve soybean seeds were 125 at CM10 and 90 at Q10 array in the mass range from 2 to 40 kDa. Among them, 82 peak clusters at CM10 and 33 peak clusters at Q10 array showed significantly different peak clusters at p<0.00004 (CM10) and p<0.00005 (Q10) among twelve different soybean cultivars. Moreover, 29 peak clusters at CM10 and 17 peak clusters at Q10 array were detected in all cultivars as an ‘universally existed peptide’. In comparison with three different agronomic types, total of 55 peak clusters (CM10) and 23 peak clusters (Q10) were significantly different peak clusters at p<0.00004 and p<0.0001, respectively. In these probability levels, soybean seeds were well discriminated into different cultivar and different type with each other. Also we could find several specific peptide biomarkers for agronomic type.

The canine major histocompatibility complex (MHC) is referred to dog leukocyte antigens (DLA), which is known to be the most polymorphic genetic system in canine species. Many cloned dogs have been produced since Snuppy, first cloned dog, there was no research about genetic identity of MHC among cloned animals. Recently in Lee’s group, two non-transgenic cloned beagles (BG1, 2) were produced by somatic cell nuclear transfer (SCNT) using fetal fibroblast (BF). Also, four transgenic cloned beagles (Ruppy 1-3, 5) were generated using transgenic BF transfected with Red fluorescent protein (RFP) gene. We hypothesize that non-transgenic (BG1, 2) and transgenic (Ruppy 1-3, 5) cloned beagles derived from identical donor cells have the same immunological genetic characteristic except for RFP gene insertion in the genome. Thus, the aim of this study is to confirm the immunological identity of DLA class II in cloned beagles produced using same nuclear donor cell. Genomic DNA was extracted from blood of BG1, BG2, Ruppy 1, 2, 3 and 5. Genomic DNA of normal two control beagle, no correlation with BF was also investigated for rulling out the possibility that beagles were inbred. Forward and reverse primers used for DLA-DQA1 and DQB1 respectively were DQAF: 5’-TAAGGTTCTTTTCTCCCTCT-3’ and DQAR: 5’-GGACAGATTCAGTGAAGAGA-3’ DQBR:5’-CTCACTGGCCCGGCTGTCTC-3’ and DQBR: 5’-CACCTCGC CGCTGCAACGTG-3’. Polymerase Chain Reaction (PCR) products were purified, sequenced directly using the Big Dye Terminator kit. Sequencing analysis was performed on an automated 3730xl DNA analyzer. In experiment 1, sequence of DLA-DQ alpha 1 (DQA1) and DLA-DQ beta 1 (DQB1) exon 2, hypervariabel region, was compared in BG1 and BG2. Experiment 2 also compared the sequence of DQA1 and DQB1 among Ruppy 1, 2, 3 and 5. Experimental 3 compared sequence of DQA1 and DQB1 among all cloned dogs (BG1, BG2 and Ruppy 1-3, 5). As a result, BG1 and BG2 have same allele for DQA1 and DQB1 as we expected. They share DQA1*00101 and DQB1*02901 in experiment 1. In experiment 2, Ruppy 1, 2, 3 and 5 also have identical DQA1*00101 and DQB1*02901 allele. No discrimination between transgenic dogs and cloned dogs was seen in DQA1 and DQB1 Allele in experiment 3. DQA1, DQB1 allele was identified as *00101 and *02901 in all dogs. We provided the allele identity of DQA1and DQB1 in cloned beagles, which can be used as preliminary data for immunological related studies. In conclusion, transgenic cloned dogs despite of red fluorescent protein genes being inserted in their nuclear DNA were immunologically compatible with non-transgenic cloned dogs. We demonstrated that cloned beagles produced using identical nuclear donor were immunologically compatible.