초록 ▼

Ⅳ. 연구개발결과
○ 1세부 : 농가 보유 4계통 재래닭 집단유전학적 특성평가
- 농가로부터 연산오계, 현인흑계, 황봉, 횡성약닭에 대한 종란 수집 및 부화, 사육 실시
- 계통별 외형조사 실시 : 성장률, 깃털색, 부리색, 정강이색, 볏색, 깃털무늬 조사
- 계통별 경제형질 조사 : 계통별 체중조사 및 20주령 이후 암컷에 대해 산란률 조사
- 재래닭 원시생식세포주 확립 : HH28단계의 배아로부터 원시생식세포 분리 후 체외배양
- 생식선키메라 확립 : 확립된 재래닭 원시생식세포를 미세주입하여 키메

Ⅳ. 연구개발결과
○ 1세부 : 농가 보유 4계통 재래닭 집단유전학적 특성평가
- 농가로부터 연산오계, 현인흑계, 황봉, 횡성약닭에 대한 종란 수집 및 부화, 사육 실시
- 계통별 외형조사 실시 : 성장률, 깃털색, 부리색, 정강이색, 볏색, 깃털무늬 조사
- 계통별 경제형질 조사 : 계통별 체중조사 및 20주령 이후 암컷에 대해 산란률 조사
- 재래닭 원시생식세포주 확립 : HH28단계의 배아로부터 원시생식세포 분리 후 체외배양
- 생식선키메라 확립 : 확립된 재래닭 원시생식세포를 미세주입하여 키메라 생산

○ 2세부 : 농가 보유 4계통 재래닭 분자유전학적 특성평가
- 재래닭 total RNA bank 및 유전체 발현 DB구축
- 재래닭 계통별 특이발현 유전자 발굴(93개) 및 표현형과의 연관관계 분석
- 재래닭 계통 특이적 마커유전자 선별(35개) 및 SNP 분석
- 효모세포 시스템 기반 멜라닌 효능 평가 : Mutant stain 생산 및 CLS 측정, ROS 측정
- 산업재산권 출원 : (1) 닭 품종 판별용 조성물 (10-2015-0123559), (2) 연산오계 판별용 조성물 (10-2015-0123544)

○ 1협동 : 농가 보유 4계통 재래닭 도체특성 및 영양, 유효성분 평가
- 재래닭 육질특성 평가 결과, 재래닭은 백색레그혼에 비해 조직이 단단하고 보수력이 낮은 특징을 보였고 재래종간에도 조직감, 지방산 조성(n6/n3 비율), 육색, 관능검사에서 차이가 있었음
- 재래닭의 저장중 산화특성 평가 결과, 고기 부분에서 SOD, GSH-Px 및 유리철은 지방산화를 촉진또는 억제하는 역할을 하는 것으로 보임
- 재래닭의 영양학적 성분 분석 결과, 연산오계의 다리육 및 가슴육에서 지방과 칼륨, 필수 지방산 함량이 높았고, 횡성약닭 가슴육에서 지방, 칼륨, 다리육에서 인, 비타민 A, B1, B3, 포화지방산 함량이 유의하게 높았으나 불포화지방산 함량은 유의하게 낮았음. 특히 정미에 가장큰 영향을 미치는 methionine과 glutamic acid 함량이 다른 품종에 비해 현저히 높았음. 현인 흑계는 콜레스테롤, 칼륨, 인, arachidonic acid, proline 및 cysteine 향량이 다른종에 비해 높았음. 황봉의 경우 다리육에서 인과 비타민 A의 함량이 높게 나왔음.
- 재래닭의 약리학적 특성 평가 결과, 모든 재래닭 시료에서 대식세포의 증식과 식균작용의 증가, IL-4 발현 감소가 확인되었음. 또한 성장기 흰쥐의 장골 길이 성장을 통계적으로 유의하게 증가 시켰음. 성장 실험 결과 성장판 길이 증식 효능 역시 확인함. 이를 통해 외래종 대비 재래닭의 영양학적 및 약리학적 효과를 확인함

○ 2협동 : 농가 보유 4계통 재래닭의 고문헌 연구 및 디지털 콘텐츠화
- 고문헌에 대한 기초조사 및 재래닭 관련 원본 확보(총 41종 74책)
- 고문헌의 번역 및 국역화, 용어 표준화, 정보 온톨로지 구축
- 고의서의 재래닭에 관한 문헌 내용 디지털 콘텐츠화를 통한 DB 구축 및 software 개발

○ 3협동 : 농가보유 재래닭의 수집 및 증식에 관한연구
- 자원의 교류방안 및 시설 계사의 소독방법 제시
- 난계대 질병억제프로그램 적용방법 제시
- 화염소독, 물리적 소독을 병행한 소독 방법 제시 등
- 농가보유 자원의 원산지 확인방안 제시
- DNA 프로파일을 이용한 혈연관계 추적방안 마련

Abstract ▼

1. A study on appearance, economic traits and preservation system of korean native chickens (PJ01053301)
Ogye, Hyunin Black, Hoengseong Yakdak, Hwangbong are korean native chicken and have bred with distinct appearance from other breeds. These breeds have be bred by units of farm and there are no

1. A study on appearance, economic traits and preservation system of korean native chickens (PJ01053301)
Ogye, Hyunin Black, Hoengseong Yakdak, Hwangbong are korean native chicken and have bred with distinct appearance from other breeds. These breeds have be bred by units of farm and there are no preservation system and classification system according to economic traits and breed. In this study, we check the appearance and measure the economic traits of each korean native chicken. Also, we tried to establish preservation system for each breed using primordial germ cells (PGCs). As results, we summarized the basic features such as feather color, chest size, chest width, tail length, thigh length and leg length of each breed with sex. Also, we check the economic traits such as body weight and egg laying rate. For establishment of preservation system for breed, we used PGCs as genetic resource. We established PGC lines of each breed and analyzed expression pattern and migration ability to confirm the pluripotency and germline competency of established PGCs. finally, we produce the germline chimera using korean native chicken. These results will be applied to establishment of preservation system of korean native chicken. Also, these data will serve as standard for selection and improvement of korean native chicken according to economic traits.

2. A study on the genetic characteristics of Korean native chicken (PJ01053302)
To identify genetic characteristics of Korean native chickens, this study was measured transriptome, and explored SNP variances on their promoter region. For the transcriptome mesurement, we collected liver samples from 13 weeks old juvenile female chicks in Ogye, Hyunin Black, Hoengseong Yakdak and Hwangbong. Then we extracted total RNA following to the protocol of TRIzol Reagent from ThermoScientific corp. with DNase I treatment. The extracted total RNA collection was verified on both spectrophotometer and formaldehyde agarose (FA) gel electrophoresis. To construct transcriptome database, we prepared target samples following to the protocol of Affymetrix^Ⓡ WT PLUS Amplification and Labeling using GeneChip^Ⓡ Chicken Gene ST array. After array images were digitalized using Software programs, Affymetrix^Ⓡ GeneChip^Ⓡ Commend Console (AGCC) Portal and Affymetrix^Ⓡ GeneChip^Ⓡ Expression Console, we performed assessment of array results. All array files were satisfied good quality, and we normalized and construct chicken transcriptome database by quantile-normalization algorithm, RMA.
To select differentially expressed genes (DEGs) for Ogye, Hyunin Black, Hoengseong Yakdak and Hwangbong, we calculated one-way ANOVA and fold-change (FC) values and filtered by statistical criteria (p-value <0.05 and FC >1.5). In the result, we found 92 genes as DEGs. Then we varified strain specific gene expression by principal component analysis (PCA). By Tukey’s post hoc test, we could selected 35 strain specific genes as genetic marker. Among the 35 marker genes, 28 genes were technically verified using qPCR. We found high correlation (0.75) between microarray signal intensity and qPCR CT values.
To identify marker gene expression regulating SNPs, we sequenced and compared promoter region of the marker genes. First, we extracted genomic DNA using proteinase K and RNase A to liver samples. Then we could prepare genomic DNA library for all Korean native chickens. After PCR amplification for specific to promoter region of the LOC768589 marker gene, we sequenced and aligned to discover SNPs. As a result, we could find Hwangbong-specific SNP at 9182903 stie, which is annotated by NCBI nuccore database. To evaluate melanin effect, we established cell evaluation system using yeast. We tried to treat melanin in two different ways; (1) direct addition of melanin as a powder form to yeast media, and (2) melanin as solution form to yeast media. As results, the first way showed extra melanin as precipitated surplus. We measured yeast growth curve and lifespan using second way. And we generated superoxide dismutase deleted strains, sod1Δ and sod2Δ mutant yeast to establish cell evaluation system of melanin effect. Under three different concentration (saturation, 1/10 dilution and 1/100 dilution) of melanin, wild, sod1Δ and sod2Δ yeast did not show significantly different growth curves and lifespan. In addition, although we measured ROS generation in each condition, no significant changes were found, but mutation effect were displayed.

3. A study on the quality and nutritional characteristics of Korean native chicken (PJ01053303)
This study was conducted to compare carcass yield, meat quality, nutriitional value and active ingredients among four lines of Korean native chickens (Yeonsan Ogye, Hyunin Black, Hoengseong Yakdak and Hwangbong) and White Leghorn as control. Chickens (N=23, male) were grown under same conditions and slaughtered at 56 weeks old to observe the physicochemical differences in breast and leg meats. The live and carcass weights of Hwangbong were significantly higher than other chickens. Four lines of Korean native chickens, regardless of the part, had higher shear force value than White Leghorn. In addition, the breast meat of Korean native chickens had higher cooking loss and lower water holding capacity than that of White Leghorn. The cooked meat of Hwangbong particularly were more chewy, gummy and cohesive with lower tenderness than other chickens. Oleic acid content which is related to meat flavor was significantly higher in the breast meat of Hoengseong Yakdak (39.6%). The meat of Korean native chickens had lower n6/n3 ratio, in which the breast and leg meat of Hoengseong Yakdak contained the lowest ratio. The meat of Hwangbong and Hoengseong Yakdak were more acceptable than the others according to sensory test.
The result of observe the association between antioxidant enzyme activity, free iron content and lipid oxidation of Korean native chicken (KNC) meat during refrigerated storage. Four lines of KNC (Yeonsan ogye, Hyunin black, Hoengseong yakdak and Hwangbong) were raised under similar conditions. A total of 16 roosters were randomly sampled and slaughtered at the age of 12 mon. The breast and thigh meats were stored aerobically for 10 d at 4°C. Although thigh meat had higher antioxidant enzyme activity, it was more susceptible to lipid oxidation and released more iron during storage than breast meat. Aerobic refrigerated storage for 10 d significantly decreased the activity of antioxidant enzymes and increased the amount of free iron and malondialdehyde. The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were negatively correlated with lipid oxidation, whereas that of catalase was not. The amount of free iron was positively associated with lipid oxidation. We concluded that chicken line did not affect strongly on antioxidant enzyme activity and lipid oxidation in breast meat of KNC. However, the thigh meat of Hwangbong and Hyunin black had higher SOD and GSH-Px activity, respectively, and lower malondialdehyde contents than that of other chickens. SOD, GSHPx and free iron play significant roles in meat lipid oxidation during refrigerated storage.
The results of the nutritional analysis of the 4 Korean native chicken (KNC) showed that lipid and potassium content was high in breast and thigh meats of Yeonsan ogye. The essential amino acid content was highest in Yeonsan ogye compared with other KNC. Hoengseong yak dak contained high content of lipid and potassium in breast meat, and high content of phosphorus, vit A, vit B1, and B3 in thigh meat. Saturated fatty acids (FA), palmitate and stearate, contents were significantly high and poly unsaturated FA content was low in Hoengseong yak dak. Bitter taste amino acid (AA), (valine, methionine, proline) and good taste AA (glutamate, methionine) contents were significantly high compared with other KNC. Furthermore, bitter taste AA (isoleucine, histidine, lysine, tyrosine) contents were significantly lower than other KNC. Hyunin black showed high contents of cholesterol, potassium in both breast and thigh meat. Arachidonic acid, proline, and cysteine contents were significantly higher than other KNC. Hwangbong contained high vit A and potassium (thigh) contents.
The analysis of functional characteristics of KNC meat revealed that all lines of KNC increased macrophage proliferation and phagocytosis, and inhibited the IL-4 expression. Furthermore, the longitudinal bone growth of adolescent rat was significantly increased. The expressions of BMP-2 and IGF-1 were also increased.
Yeonsan ogye increased the proliferation of osteoblasts, ALP activity, and growth plate height. Hoengseong yak dak increased ALP activity, inhibited NO secretion and IL-10 expression. Hyunin black increased ALP activity and inhibited NO secretion in in vitro analysis, and increased growth plate height in in vivo analysis. Hwangbong increased osteoblasts proliferation and inhibited NO and IL-10 expressions.

4. Analysis and digitalize of the old document for Korean native chicken (PJ01053304)
This study analyzes the chicken-related prescription shown in the literature summarized by standing tradition deliberate aims to investigate the clinacal efficacy of native chickens. Also, this study is to establish the analysis and separated by organized to establish a database on the basis of prescription and mixed herbs in a variety of functional food developed through usability evaluation methods of treatment.
Domestic Research Status in the native chicken has announced the physiological activity results for the cosmetic effect, blood purification, cerebral thrombosis, angina pectoris, hypertension, hepatitis, kidney disease, are reported to be useful in the treatment of fractures, the antimicrobial activity of lysozyme separated from the egg white of eggs of Yeonsan ogye. However, it made for the industrialized material to clinical use of the musculoskeletal system and endocrine and circulatory diseases. In this results, chicken items were produced research and development of food supplements.
Retention based investigation and farmers of the literature related to the four strains were obtained continuously source of native chickens. And The old documents were translated and build the ontology was built in traditional medicinal systems for chicken recipe. Also, documents were medical information extraction and data processing and database designs. Finally, Native chicken of Four strains were produced development based composition through digital contents that were developed software.
The management systems of native chickens related to lack of knowledge and information to foster the continued expansion and specialized training and knowledge of the future growth engine industry through the added value of cultural projects of national knowledge resources. Finally, convergence research and evaluate the usefulness of native chickens through interdisciplinary research systems, and practical measures of functional foods and alternative medicine are presented as well.

5. A study on collection and proliferation of Korean native chicken (PJ01053305)
Chicken industry of Korean Livestock sector is most biggest potion as an economically view. But, now day of Chicken industry in Korea, mainly depending on Exotic breeds. Aims of these studies are focusing for development of new breeding materials form native chicken which are keeping in small scale framers in Korea. Total of 1030 eggs are using for hatching, 430 chick are hatching from 1030 ferilazied eggs. Before hatching all eggs are disinfection with normal method for prevention of contagiousness disease and perforation in edge of eggs to injection antibiotics for encourage of early growth. We are also development method to prevention of epidemic diseases using flame.
Total of 281 DNA samples belong 5 population are used for MS marker analyzing. 30 MS markers are mobilzed of molecular genetic research. Highest allele composition population is Yeonsan Oke, that was showed 155 allele. Yatak are showed lowest allele composition, it was 41 allele in 23 MS marker. Genetic variation was estimated by Averager Heterozygous(H) degrees. The values of H in Hyunin Black and Oke populations are shown the higher than Leghorn breeds, But H values of Hwangbong and Yatka are shown lower then leghorn breed. It seems that origins of these population was very small. Coidentity of population are calculated by Structure program. Oke population are divide by two sub population. Genetic relationship of these Hwangbong, Hyunin Black and Yatak are very closer but Oke is little be far from these population.
Total of 59 DNA samples belong 5 population are used for analyzing of full length of Mitochondria DNA. Total length of Mitochondria DNA are composition of 16,780bp. With in these length 119 positons are shown segregating site. Total of 45 haplotype are discovery on these samples and 36 kind haplotypes are only shown in Korean native chicken. Genetic relationship which are calculated by individual base pair sharing distance indicate that HwangBong and Yaktak are clustering as an independent same groups, But Okey and Hyunin Black is sprayed 3 cluster.
These results are strong suggesting that these 4 Korean native chicken populations are independently rising and grow up with very limted gene exchange. Also these population need to more deep investigating.