In this study, we report for the first time a hybridization between Caretta caretta and Chelonia mydas in South Korea. On 7th May 2012, a sea turtle (Jeju sea turtle 01, JST01) was found in the east coast of Jeju Island, South Korea. The morphological characteristics of JST01 were compared with those of C. caretta and C. mydas that were frequently observed in Korean Seas. Interestingly, JST01 showed similar morphological characters such as body color, appearance of carapace, shape of upper beak, serration on marginal scute, and two claws with those of C. caretta. On the other hand, JST01 showed also similar patterns in several characteristics including the numbers of marginal, inframarginal and costal scute, smooth carapace without keel, no contact between costal scute and nuchal scute with those of C. mydas. From these results, we suggest that JST01 is one of hybrid animals bred between C. caretta and C. mydas, because JST01 has speciesspecific characters from both species. We anticipate that our study would be useful and have significant impact onto study inter-specific hybridization in the groups of sea turtle.

Precise, rapid and simple methods for species identification in animals are among the most important techniques in the livestock industry and research fields including meat classification. In this study, polymerase chain reaction (PCR) based molecular identification using inter species polymorphisms were examined by PCR-restriction fragment length polymorphism (RFLP) analysis for mitochondrial DNA (mtDNA) cytochrome b (CYTB) gene sequences among four mammalian livestock animals (cattle, horse, goat and pig). The results from PCR-RFLP analysis using the AluI restriction enzyme were also provided for the species-specific band patterns among CYTB gene sequences in these four species. The AluI-digestion for CYTB genes provided interesting migration patterns differentially displayed according to each species. Cattle and horse had one AluI-recognition site at different nucleotide positions and their AluI-digested fragments showed different band patterns on the gels. Pig had two AluI-recognition sites within the amplified CYTB sequences and produced three bands on the gels. Goat had no AluI-recognition site and was located at the same position as the uncut PCR product. The results showed the species-specific band patterns on a single gel among the four livestock animal species by AluI-RFLP. In addition, the results from blind tests for the meat samples collected from providers without any records showed the identical information on the species recorded by observing their phenotypes before slaughter. The application of this PCR-RFLP method can be useful and provide rapid, simple, and clear information regarding species identification for various tissue samples originating from tested livestock species.

한국산 집쥐속(Rattus)에 속하는 동물들을 2005년 4월부터 2012년 10월까지 제주도의 각처에서 채집하여 형태학적, 유전학적 종 동정과 형태적 특성, 2차 성적이형, 서식지 유형 등을 조사하였다. 연구결과, 제주도에서 포획된 Rattus 속 야생쥐들은 Rattus norvegicus와 R. tanezumi로 확인되었고, R. rattus는 확인되지 않았다. R. norvegicus는 제주도내 각처에서 특히, 주택 밀집지역의 인가 내-외부에서 채집되었고, 일부는 자연환경인 해안이나 경작지, 산림에서 관찰되기도 하였다. R. tanezumi는 동물 사육 농장의 내외부에서만 확인되었다. 형태적 특성으로 R. norvegicus는 꼬리길이(TL)와 귀의 길이(EL)가 R. tanezumi에 비해 짧고, 이들 형질의 머리-몸통 길이의 비율인 T_HR, E_HR 등도 유의적인 차이를 보였다(p<0.05). 인구밀집 지역인 도심과 농촌 지역의 R. norvegicus와 개체군 사이에서는 도심지역 개체군의 체중이 더 무거운 것으로 확인되었다(p<0.05). R. norvegicus에서는 2차 성적이형 현상이 나타나지 않았으나, R. tanezumi는 암컷이 수컷에 비해 더 무거운 것으로 나타났다(p<0.05). 현재까지 보고된 연구결과와는 다소 다른 종의 분포와 서식환경을 보여주고 있는 본 연구 결과는 향후 추가연구를 통해 제주도와 한반도 포유류상의 재검토와 함께 서식지 분포 양상과 생태적 특성 등을 명확하게 구명해야함을 시사하고 있다.

This study was carried out to investigate synthetic extender for semen cryopreservation of Jeju Native Black Bull. The semen was collected using an artificial vagina and transported to the laboratory. The semen was diluted 1:1 by Tris-Egg yolk extender and contrifuged in 1,500 rpm for 15 minutes. The supernatant was removed. The pellect was diluted to final sperm concentration of 2×108/ml by doubling in every 30 minutes at 4℃ cold chamber. The semen was equilibrated for 4 hours at cold chamber and packed to 0.5 ml straw. The semen straws were located above 5 cm for 10 minutes. The height and duration affect the freezing speed by temperature. The frozen straw was plunged to LN2. The presented straws were examined the viability and motility after thawed at 37℃ water bath. Frozen-thawed sperm were evaluated sperm viability, membrane integrity and acrosome integrity. Post-thawed sperm viability has been significantly higher (p<0.05) in fresh sperm (93.27±1.62%) than frozen-thawed sperm (73.34±3.27%). However, there were no significant differences between fresh and frozen-thawed dead cell rate (7.35±2.63 vs, 13.71±2.85). In sperm motility, between Triladyl and AndroMed Extender, there was no significant different (72.86±2.83 vs, 81.47±2.48), similarly, the dead cell rates was similar (18.41±3.42% and 17.26±4.25). The results of our study suggest that AndroMed to the freezing extender showed more positive effect on the frozen-thawed spermatozoa in Jeju Native Black bull semen.

본 연구는 경상북도 청도군 운문산 자연휴식년제 지역일대의 포유류상을 밝히기 위해 2007년 12월부터 2009년 9월까지 수행되었다. 조사결과 6목 11과 24종의 포유류의 서식을 확인하였다. 우점종으로는 고라니 Hydropotes inermis, 다람쥐 Tamias sibiricus, 두더지 Mogera wogura, 흰넓적다리붉은쥐 Apodemus peninsulae 등이었다. 특히, 천연기념물(NM)과 환경부지정 멸종위기종(ES)인 하늘다람쥐(Pteromys volans; NM #328, ES II급), 수달(Lutra lutra; NM #330, ES I급), 삵 (Prionailurus bengalensis; ES II급), 담비 (Martes flavigula; ES II급) 등 법적보호종 4종이 발견되었다. 다양한 포유류 종과 법적보호종의 서식에 대한 지리적 분포를 확인한 이 결과는 조사지역이 포유류의 서식에 필요한 자연생태계의 매우 양호한 조건을 갖추고 있는 지역임을 반영한다. 반면, 관리동물 종으로 지정된 들고양이가 다수 관찰되었다. 운문산 자연 생태계의 건전한 유지와 생태계 위해 동물의 관리 방안을 마련하기 위해서는 지속적이고 장기적인 연구조사가 필요하다.

The present study was to assess the in vitro viability and sexing rate of bovine embryos. Blastocysts were harvested on day 7～9 day after insemination(in vitro and in vivo), and the sex of the embryos was examined using the LAMP method. Embryo cell biopsy was carried out in a 80 μl drop Ca2+, Mg2+ free D-PBS and, biopsied embryos viability were evaluated after more 12 h culture in IVMD culture medium. The formation of recovered embryo to expanded and hatching stages had ensued in higher of sexed embryo in vivo than in vitro (100% vs. 89%, p<0.05), and in vitro, the rates of degeneration after sexing were significantly (p<0.05) higher in vitro than in vivo(11% vs. 0.0%). The rates of the predicted sex were female 61% vs. 56%, and male 39% vs. 44% in vivo and in vitro, respectively. The rates of survival following different biopsy methods were seen between punching and bisection method in vivo and in vitro (100% vs. 89% and 100% vs, 78% respectively). Biopsy method by punching was significantly (p<0.05) higher than bisection between produced embryos in vivo and in vitro. The present data indicate that with microblade after punching for embryo sexing results in high incidence of survivability on development after embryo biopsy. It is also suggested that LAMP-based embryo sexing suitable for field applications.

Oxidative stress has been reported to be one of causes of neuritis. This study examined antioxidative activities of methanol extracts of six amphibian species known to be medicinal animals (Rana catesbeiana, R. coreana, R. rugosa, R. dybowskii, R. nigromaculata, and Hyla japonica) and investigated their effects of inhibiting nitric oxide (NO) production and cytotoxicity on the murine macrophage RAW264.7 cells. As inflammation is closely associated with reactive oxygen species, assays on 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity, xanthine oxidase inhibitory activity, superoxide anion radical scavenging activity and NO scavenging activity of the extracts of the six species were performed to investigate their antioxidative activity. The results obtained were as follows; All extracts showed antioxidative activity, and the activity of R. dybowskii was the highest in comparison among those. Anti-inflammatory effects of the extracts were also examined, the five extracts except that of R. rugosa did not show cytotoxicity for RAW264.7 cells at the maximal concentration (1,000 μg mL-1). Selectivity index, meaning NO scavenging activity compared to cytotoxicity, showed the highest level in the extract of R. dybowskii. These results will be very useful basic data for future studies on prevention and treatment of human diseases to understand the biological roles of amphibian extracts throughout the antioxidative or anti-inflammatory pathways.

The aim of this study was to evaluate the effect of α-tocopherol on blastocysts development and subsequent cryosurvival of the vitrification. The α-tocopherol(0, 100, 200, 400 μM) was added in to culture medium for the bovine embryos. The blasocysts from the α-tocopherol and untreated control groups were then frozen-thawed, and their cryosurvival was assessed by in vitro culture for 48 h. There were no differences in the overall cleavage rate(56.14±4.66, 58.18±4.70, 62.97±6.86 and 51.17±7.28) among four treatment groups. However, in blastocyst development and total cell number were significantly higher in α-tocopherol 200 μM(38.60±7.12; 106.33±3.50) to culture medium than other treatment groups(29.30±5.24, 31.60±7.12 and 26.37±4.18; 101.36±5.12, 97.27±2.87, and 91.23±7.52 respectively). Before and after vitrification, the total cell number and blastocyst development of embryo were significantly higher in July to August than September to October. In conclusion, addition of α-tocopherol 200 μM to in vitro bovine embryo culture medium was beneficial for improving embryo quality by decreasing the embryo damage blsstocysts cell number and improving the tolerance of the embryos to cryopreservation.

This study carried out to examine the association between the haplotypes of mitochondrial control region (CR) and growth traits of two F1 progeny populations of the Red-spotted grouper. We found polymorphic patterns of the 133-bp repeat units in the CR. In the BBR01 population (70-days after hatching), a total of 1,091 F1 progeny were divided into three haplotypes (H01, H03 and H04). The significant differences were found in the levels of BL, BW and LWI (p<0.05). The F1 animals with the haplotype H01 had greater level of BL (50.815±4.586 mm) than those of H03 (47.270±6.486 mm) and H04 (47.179±6.278 mm). The H01 F1 fishes were heavier level of BW (2.270±0.559 g) than those of H03 (1.789±0.711 g) and H04 (1.797±0.706 g). In the BBR02 population (11-months after fertilization), three haplotypes H03, H04 and H05 were detected. The significant difference was found only in the BL values among three haplotypes (p<0.05). The F1 animals with the haplotype H03 had greater level of BL (19.22±2.000 cm) than those of H04 (18.64±1.964 cm) and H05 (18.86±1.512 cm). There were no significant differences in BW and LWI among haplotypes in the BBR02 population (p>0.05). These results suggested that the mitochondrial haplotypes may affect the growth traits during early developmental stage of the Red-spotted grouper. The marker-assisted selection system for broodstock animals may be helpful in improving performance traits for aquaculture of the Red-spotted grouper.

This study analyzed the mitochondrial DNA (mtDNA) sequences of the Red-spotted grouper, Epinephelus akaara (Perciformes, Serranidae), and used for construction of molecular phylogeny and for association between maternal haplotypes and phenotypic differences of F1 progeny. This study revealed phylogenetic position of the endangered red-spotted grouper, Epinephelus akaara (Perciformes, Serranidae) based on the nucleotide sequences of complete mt genome. Complete nucleotide sequences were determined from the mt genomes of two individuals of the red-spotted grouper caught in South Korea. The mitochondrial genome had 16,795 base pairs (bp) and 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs, and a noncoding control region. The two mt genomes were highly homologous (99.71% similarity). The two mt genomes of E. akaara determined in this study were found in Clade I in the phylogenetic tree with those of E. awoara, E. fasciatomaculosus, E. sexfasciatus, E. diacanthus, E. sticus, and E. morio, suggesting that this may be helpful to understand phylogenetic position of Epinephelus species including red-spotted grouper. The genetic structure and phylogenetic relationship were investigated in the red-spotted grouper populations using the sequence polymorphisms of the cytochrome c oxidase subunit I(COI) gene and variable number of tandem repeats (VNTRs) of the control region (CR). A total of forty-one COIhaplotypes were found from 174 COIsequences from East Asia. The Jeju Island population (n=5) had four haplotypes, and the South Sea population (n=105) had twenty-five haplotypes. The Hong Kong population had nineteen haplotypes from fifty-nine COIsequences determined in this study. Among the COIhaplotypes, EAC_03 is commonly found in all populations (Jeju Island and South Sea of Korea, China, Hong Kong and Taiwan). In addition, there were four haplotypes (EAC_12, EAC_14, EAC_28 and EAC_35) also common among the populations tested in this study and collected from NCBI database. However, twenty haplotypes were specific in the Korean populations, and fifteen haplotypes were specific in the China and Hong Kong populations. The neighbor-joining (NJ) trees constructed from the phylogenetic analyses based on the polymorphisms of the COIhaplotypes showed the monophyletic branching pattern within the genus Epinephelus, indicating that the red spotted grouper populations had evolved from common maternal ancestors. Consequently, East Asian red-spotted grouper populations are maternally related at least in part, as well as sharing the same evolutionary history, and still affected by the East Asian ocean current (Kuroshio). From the haplotype analysis for mtDNA CR, we obtained VNTR polymor-phisms in all populations tested. We found five haplotypes for the CR VNTR patterns. The 133-bp repeat units were counted two to five. Using CR VNTR haplotypes, the statistical association was examined between mtDNA haplotypes and growth traits of aquafarming young fishes of the red-spotted grouper. A total of 386 F1 progeny, which were randomly selected from a progeny population produced by artificial insemination in the farm, were genotyped and statistically compared their body length (BL), body weights (BW) and length-weight indexes (LWI) at 11-months after hatching. There haplotypes H03, H04 and H05 were detected for CR in the parents and progeny populations. The significant difference was found in the BL values among three haplotypes (p<0.05). The F1 animals with haplotype H03 had freater level of BL (19.22±2.000 cm) than those of H04 (18.64±1.964 cm) and H05 (18.86±1.512 cm). There were no significant differences in BW and LWI among haplotypes (p<0.05). These results concluded that the maternal lineages affected the growth rates during early developmental stage in the red-spotted grouper. These findings suggested that the mitochondrial background of the fertilized eggs may play an important role in the early development, and the markerassisted selection system for broodstork animals may be helpful in improving performance traits for aquaculture industry as well as for conservation biology of the endangered red-spotted grouper. However, the results from the association analysis between haplotypes and phenotypes of F1 progeny (n=1,093) at 60-days after hatching showed that there were no significant difference (p>0.05). Consequently, the results of this study may be useful information for understanding the evolutionary relation with other species and may be good genetic markers for breeding management in the red-spotted grouper aquaculture system.

As a preliminary investigation into the effect of environmental factors control for gonadal development, we examined the involvement of photoperiod and water temperature in ovarian development of Epinephelus. akaara. For the induction of sexual maturation, E. akaara reared in recirculating aquaculture system (RAS). During November 2013, the photoperiod and water temperature was adjusted to 12L:12D and 18℃, respectively. In the photo-thermal treatment group, every 3 weeks daylight was increased as follows a 13L:11D and 14L:10D, and control group was maintained under natural condition. After 9 weeks, water temperature was increased 23℃ in photo-thermal treatment group. The sampled fish every 3 weeks revealed increase in gonadosomatic index (GSI; 5.18±1.38), oocyte diameter and vitellogenic oocytes (423.9±36.1 ㎛) were observed in gonads 12 weeks under photo-thermal treatment group. However, ovarian development was maintained immature stage in control group. In this environmental factors manipulation trial, seventy one of the 95 females (578.4 ± 25.4 g in mean body weight, 31.0 ± 0.5 cm mean total length) treated with HCG injection (doses 500 IU/kg BW) were induced ovulation by artificial stripping. The total volume of ovulated eggs were 3,470 ml and the total volume of fertilized eggs was 3,295 ml. The fertilization rate and hatching rate were 95% and 98%, respectively. These results suggest that the photoperiod as well as water temperature are major environmental factors in triggering the gonadal development of E. akaara.