전장유전체 관련성 분석을 통한 순종 랜드레이스 생시체중 형질의 위치상 후보유전자 탐색

강호찬,이재봉,유채경,최태정,임현태 경상대학교 농업생명과학연구원 2018 농업생명과학연구 Vol.52 No.3

돼지의 생시체중은 생존율과 폐사율에 밀접한 관련이 있어 양돈산업에서 자돈 관리와 직결된 중요한 경제형질이다. 본 연구는 Genome-Wide Association Study(GWAS) 분석을 통해 순종 랜드레이스의 생 시체중과 관련된 위치상 후보유전자 탐색을 실시하였다. 생시체중의 유의적 관련 분석 결과, genomewide suggestive level에서 유의성 있는 single nucleotide polymorphism(SNP) marker는 3번 염색체 (ASGA0098921, P=2.41×10-5)와 4번 염색체(H3GA0013451, P=2.47×10-5)에서 각각 1개씩 동정되었다. 이들 SNP marker가 위치한 3개의 유전자(LOC110260055, LOC100156472, LOC100157689)들은 순종 랜드레이스 생시체중의 위치상 후보 유전자이며, 이들 유전자 정보를 이용해서 순종 랜드레이스 생시 체중을 선발할 수 있는 기초 연구 자료가 될 것으로 사료된다. Birth weight represents an important trait that has an economic value because it is closely associated with survival and mortality. This study aimed to identify positional candidate genes associated with birth weight of purebred Landrace pigs using genome-wide association study(GWAS). Based on the analysis of genes significantly associated with birth weight, a single nucleotide polymorphism(SNP) marker that is significant at genome-wide suggestive level was identified on chromosome 3(ASGA0098921, P=2.41×10-5) and 4(H3GA0013451, P=2.47×10-5), respectively. These three genes(LOC110260055, LOC100156472, LOC100157689) containing each SNP marker individually are positional candidate genes associated with birth weight of purebred Landrace pigs. Therefore, the results of this study can be used for fundamental studies required to select an optimal birth weight range for the purebred Landrace pigs.

순종 랜드레이스의 10주령 체중 형질에 영향을 미치는 후보유전자 탐색을 위한 전장유전체 분석

강호찬,이재봉,유채경,최태정,임현태 강원대학교 동물자원공동연구소 2019 동물자원연구 Vol.30 No.3

According to Livestock Inspection Standards, the piglets enter the feedlot at approximately 30 kg, and the inspection starts after the preliminary feeding period. The reason for applying the preliminary feeding period is to select inspection piglets with no diseases after the complete growth of the internal organs until 10 weeks of age. Furthermore, the age of 10 week is the time when the muscle fibers grow to their maximum size and the piglets are prepared for fat deposition at the later fattening period. In the study, a genome-wide association study (GWAS) was performed through the mlma command of the genome-wide complex trait analysis (GCTA) program with 703 purebred Landrace population, and the candidate genes associated with the weight of 10 week were searched. The GWAS identified 3 single nucleotide polymorphism (SNP) markers, which have a significant genome-wide suggestive level, on chromosome 6 (DIAS0002615; p value=1.62×10-6, MARC0083933; p value=4.94×10-6, ASGA0028717; p value=5.40×10-6). The 2 genes (Ubiquitin protein ligase E3 component n-recognin 4; UBR4, WD and tetratricopeptide repeats 1; WDTC1) in which these 3 SNP markers are located are positional candidate genes of the weight of 10 week of the purebred Landrace population. 2 candidate genes have been reported to be associated with fattening. Therefore, the positional candidate genes in this study, UBR4 and WDTC1, are expected to be usable as genes for traits associated with the weight of 10 week weight and fattening through additional experimental research with other population 가축검정기준(Livestock Inspection Standards)에 따르면 약 30 kg에 입식을 하여 예비사육기간 이후 검정을 시작한다. 이는 10주령 시기까지 체내 기관의 온전한 성장과 질병에 문제가 없는 검정자돈을 선발하기 위함이며, 또한 10주령 시기는 근섬유의 크기가 최대치로 증가하고 이후 비육시기의 지방 침착을 위해 준비하는 시기이다. 이에 본 논문은 순종 랜드레이스 집단 703두와 genome-wide complex trait analysis(GCTA) 프로그램의 mlma command를 통한 전장유전체 연관성 분석(genome-wide association study, GWAS)을 수행하여 10주령 체중과 연관된 후보유전자를 탐색하였다. 전장유전체 연관성 분석 결과 genome-wide suggestive level의 유의성을 가지는 single nucleotide polymorphism(SNP) marker가 6번 염색체에서 3개 동정되었다(DIAS0002615; p value=1.62×10-6, MARC 0083933; p value=4.94×10-6, ASGA0028717; p value=5.40× 10-6). 이 3개의 SNP marker가 위치한 2개의 유전자(Ubiquitin protein ligase E3 component n-recognin 4; UBR4, WD and tetratricopeptide repeats 1; WDTC1)들은 순종 랜드레이스 집단의 10주령 체중의 위치상 후보 유전자이며, 두 개의 후보유전자는 비육과 연관된 유전자로 보고되어 있다. 따라서 본 연구의 위치상 후보유전자인 UBR4, WDTC1은 타 집단과 추가적인 실험 연구를 통해 10주령 체중과 비육 연관 형질의 유전자로서 이용이 가능할 것으로 생각된다.

Identification of marker in processed meat products of different species by using multiplex PCR

강호찬,김은호,김지영,Cheol-Hyun Myung,Jae-Bong Lee,Chae-Kyoung Yoo,임현태 한국동물유전육종학회 2021 한국동물유전육종학회지 Vol.5 No.4

A processed meat product is a food that changes the character or shape of meat, and it is difficult to identify the raw meat because the original shape is destroyed. False record and disguised sales of raw meat can negatively affect not only the safety of consumers but also the income of producers. Therefore, this study was conducted to establish a multiplex PCR (polymerase chain reaction) method to simultaneously identify cattle (Bos taurus), pigs (Sus scrofa), and chickens (Gallus gallus), which are mainly used as raw meat in the manufacture of processed meat products. As a result of species-specific PCR of the raw meat DNA to confirm the cross-reactivity, species-specific amplification products of cattle 384 bp, pigs 249 bp, and chickens 721 bp were identified, and no other non-specific amplification appeared. Based on this, in order to evaluate the applicability in the market, multiplex PCR was performed by classifying 52 processed meat products on sale by 5 types (Ham, Tteokgalbi, Pattie, Can ham, Sausage). As a result, the indicated ingredients were detected in all products except Can ham No. 7 among 52 processed meat, and specific reactions were observed in chickens that were not marked in No. 1, No. 4 of Tteokgalbi. In particular, among the pigs and chickens marked on Can ham No. 7, only pigs showed a specific reaction, confirming the possibility of false record. If this study is used to identify the types of raw meat used in processed meat products, it is expected that it will contribute to improving awareness and promoting consumption of processed meat by reducing cases of illegal distribution and restoring consumer confidence in processed meat. 본 연구는 육가공품에서 종 식별 마커의 multiplex PCR 방법에 대한 적용 가능성을 확인하고 실시하였다. 육가공품에서 가장많이 사용되는 소, 돼지, 닭의 종 식별 마커를 선별하여 원료육과 가열육에서 추출한 DNA를 활용하여 PCR을 실시하였다. 그 결과, 원료육과 가열육에서 소, 돼지, 닭의 종 식별 마커가 교차반응 없이 모두 식별이 가능하였고, 검출한계 실험은 원료육에서 소, 돼지, 닭이 최대 0.01 ng/㎕, 가열육에서는 소는 최대 0.01 ng/㎕, 돼지와 닭은 최대 0.1 ng/㎕으로 가열로 인한 DNA 파괴를 고려하더라도 육가공품에서 충분히 종 식별이 가능할 것으로 판단된다. 이후, 5종류로 나눈 52개의 육가공품에 multiplex PCR를 실시하였다. 표기된 성분과 종 식별 PCR 결과, 햄류, 패티류, 소시지류에서 표기된 성분과 multiplex PCR의 종 식별 결과가 동일하였으나, 떡갈비 1번, 4번의 제품이 표기된 성분 외에 닭의 특이적 반응이 검출됨으로써 의도적 또는 비의도적으로 닭고기가 포함되었으며, 캔 햄 7번의 제품에서 표기된 돼지, 닭 중 돼지만 특이적 반응이 검출되어 제품의 원재료명과 표기가 다른 것을 확인하였다. 본 실험을 통해 육가공품 제조 시 실제로 표기되지 않은 축종, 특히 가장 값이 저렴한 닭고기를 사용할 가능성이 있음을확인하였으며, 허위기재의 가능성 또한 확인하였다. 육가공품 원료육 표기의 정확도 향상 및 육가공품에 대한 소비촉진을 위해육가공품에 대한 원료육의 종 식별뿐만 아니라 원료육 성분비 판별을 위한 추가연구가 필요할 것으로 판단된다

Analysis of the relationship between the end weight trait and the gene ADGRL2 in purebred landrace pigs using a Genome-wide association study

강호찬,김희성,이재봉,유채경,최태정,임현태 충남대학교 농업과학연구소 2018 Korean Journal of Agricultural Science Vol.45 No.2

The overall consumption of meat is increasing as the level of national income increases. The end weight is a trait closely associated with dressed meat. Genome-wide association study (GWAS) is an effective method of analyzing genetic variation and gene identification associated with a number of natural alternative traits because it can detect variations. So this paper did a GWAS analysis to identity the location on the genome related to the end weight in purebred landrace pigs and to explore the relevant candidate gene. This study identified a significant single nucleotide poly morphism (SNP) marker in chromosome 6 (ASGA0029422, p = 1.22 x 10-6). Adhesion G protein-coupled receptor L2 (ADGRL2 ) was found to be the candidate gene at the identified SNP marker location. ADGRL2 genes have been found to be associated with cell development in relation to the external and internal environment of a cell. In addition, genotype and statistical analyses were done on nine variations on the exon of ADGRL2 . The results show that the SNP marker (ASGA0029422, p = 1.32 × 10-6) was significant, but the significance of the nine variations on the ADGRL2 exon was not verified. However, by performing further experiments and functional studies on other SNPs showing possible genetic ADGRL-Exon mutations, objects with high associations of high-end weights can be selected.

Genetic diversity and relationship analyses of the Korea native black goat line using microsatellite markers

강호찬,김관우,김은호,명철현,이정규,임현태 충남대학교 농업과학연구소 2021 Korean Journal of Agricultural Science Vol.48 No.4

The aim of this study was to analyze the genetic diversity and distance of the Korean native black goat line. Thus far, this Korean native black goat line has not been studied intensively, especially in genetic diversity and relationship studies in comparison with other breeds. In total, eleven microsatellite (MS) markers were used to evaluate alleles from 391 Korean native black goats and foreign hybrid animals. The genetic diversity index was evaluated based on the allele distributions. Four Korean native black goat lines showed expected ranges of observed heterozygosity, expected heterozygosity, and polymorphism information content (PIC) values for use in genetic diversity research (0.509 - 0.643, 0.434 - 0.623 and 0.356 - 0.567). Lines from the Korean native black goat and foreign hybrid were clearly separated according to principal coordinates analysis (PCoA), phylogenetic tree and tended to be clustered in each Korean native black goat line. Thus, this study can be used for analyzing the genetic relationships between Korean native black goats and foreign breeds for line preservation and for fundamental information to determine breed improvement strategies.

Genome-wide association study identifies positional candidate genes affecting back fat thickness trait in pigs

이재봉,강호찬,김은호,김윤주,유채경,최태정,임현태 충남대학교 농업과학연구소 2018 Korean Journal of Agricultural Science Vol.45 No.4

This study was done to search for positional candidate genes associated with the back fat thickness trait using a Genome-Wide Association Study (GWAS) in purebred Yorkshires (N = 1755). Genotype and phenotype analyses were done for 1,642 samples. As a result of the associations with back fat thickness using the Gemma program (ver. 0.93), when the genome-wide suggestive threshold was determined using the Bonferroni method (p = 1.61 × 10-5), the single nucleotide polymorphism (SNP) markers with suggestive significance were identified in 1 SNP marker on chromosome 2 (MARC0053928; p = 3.65 × 10-6), 2 SNP markers on chromosome 14 (ALGA0083078; p = 7.85 × 10-6, INRA0048453; p = 1.27 × 10-5), and 1 SNP marker on chromosome 18 (ALGA0120564; p = 1.44 × 10-5). We could select positional candidate genes (KCNQ1, DOCK1, LOC106506151, and LOC110257583), located close to the SNP markers. Among these, we identified a potassium voltage-gated channel subfamily Q member gene (KCNQ1) and the dedicator of cytokinesis 1 (DOCK1) gene associated with obesity and Type-2 diabetes. The SNPs and haplotypes of the KCNQ1 and DOCK1 genes can contribute to understanding the genetic structure of back fat thickness. Additionally, it may provide basic data regarding marker assisted selection for a meat quality trait in pigs.

Genome-wide association study for ethological traits of purebred Landrace and Yorkshire populations

최태정,강호찬,이재봉,유채경,김은호,임신재,임현태 경상국립대학교 농업생명과학연구원 2020 농업생명과학연구 Vol.54 No.2

Some behaviors of pigs that are not expressed in the wild state or are observed in a small minority of individuals after groups of pigs are mixed have been reported to indicate poor welfare. A GWAS analysis was performed by measuring the frequency and duration of the four ethological traits and using the mlma command provided by the genome-wide complex trait analysis (GCTA). The positional candidate genes on significantly identified single nucleotide polymorphism (SNP) markers were identified by using the dbSNP provided by the National Center for Biotechnology Information (NCBI). When the GWAS analysis was applied the 43,565 (of the purebred Landrace population) and 41,700 (of the purebred Yorkshire population) SNP markers, 1, 2, and 1 significant SNP markers were identified for the traits of feeding frequency (LOC110262254), locomotion time (LOC110260361), and locomotion frequency (LOC110260361) of the purebred Landrace population, respectively. Meanwhile, 1 and 7 significant SNP markers were identified for the traits of drinking time (LOC110260090) and feeding frequency (MAP3K19; LOC110257013; ACMSD; TMEM163; RAB3GAP1) of the purebred Yorkshire population, respectively. The results of this study may suggest that the GWAS analysis of the ethological traits of purebred Landrace and Yorkshire populations could be used to perform a GWAS analysis on non-economic traits, and the results can thus be provided as basic data for GWAS analyses of other non-economic traits in the future.

Identification of marker in processed meat products of different species by using Real-time PCR

김은호,강호찬,김지영,Cheol-Hyun Myung,Jae-Bong Lee,Chae-Kyoung Yoo,임현태 한국동물유전육종학회 2021 한국동물유전육종학회지 Vol.5 No.4

In order to secure consumer safety against false record and inaccurate information on processed meat products, it is necessary to establish a detection method for raw meat used in processed meat products. This study performed a quantitative analysis of raw meat of cattle (Bos taurus), pigs (Sus scrofa), and chickens (Gallus gallus) used in the production of processed meat products using the Real-time PCR method, which can confirm gene expression and detection amount in real time. For the species-specific primer design, the sequence of the D-loop and Cytochrome b (Cyt-b) gene in mitochondrial DNA (mtDNA), which are widely used for species identification due to its high mutation rate, were used. As a result of the species-specific real-time PCR, the cycle quantification (Cq) value expressed according to the species was found to be high in the order of pigs, cattle, and chickens, and no cross-reaction appeared. Based on the results of confirming species specificity, 52 processed meat products on sale were classified into 5 types (Ham, Tteokgalbi, Pattie, Can ham, Sausage), and real-time PCR was performed. All ingredients label in the processed meat products except for Can ham No. 7 were detected, and those that did not match the label were No. 2 and No. 5 in Ham, No. 2, No. 6, No. 16 in Sausage, and No. 1 and 4 in Tteokgalbi. A total of 7 products showed a specific reaction of chickens. In the case of products with different detection results from the labeled ingredients, intentional or unintentional mixing could be suspected through the difference in Cq values of the main ingredient and unlabeled ingredient. If a quantitative analysis method for raw meat is established and commercialized by optimizing these Real-time PCR conditions, it is thought that consumers' awareness and safety will be improved. 이번 연구를 통하여 육가공품에서의 종 식별 마커의 적용 가능성을 확인하였으며, 총 5종류 52개의 육가공품을 대상으로 Realtime PCR을 수행한 결과로 캔 햄 7번을 제외하고 표기된 성분이 모두 검출이 되었으며, 제품 표기와 다른 성분이 있는 제품 7가지를 확인하였다. 이를 통해 미표기된 원료육이 혼합될 가능성을 확인할 수 있었다. 현재 식품 공정에 관한 법령의 경우 육가공품의 제조 기준과 포장재 표기 사항 등의 내용은 기재되어 있지만, 원료 성분을 확인할 분석방법에 관한 규정은 없는 실정이다. 따라서, 이러한 검사법을 통해 최종 육가공품의 원료 종류 및 함량을 분석할 수 있는 기술이 필요한 것으로 보이며, 소비자 관점에서 동일한 유형의 육가공품에 대하여 사용한 원료육의 품질구분이 어렵다는 점을 고려했을 때 고기 함량 표기에 세부사항까지표기할 수 있도록 하는 규제가 필요한 것으로 사료된다.

한우 혈통 정확도 향상을 위한 SNP 정보 기반 친자 확인 평가

김지영,김은호,강호찬,명철현,임현태 경상국립대학교 농업생명과학연구원 2023 농업생명과학연구 Vol.57 No.5

본 연구는 국내의 한우 개량에 있어서 정확한 혈통 정보가 중요해짐에 따라 고밀도 Single Nucleotide Polymorphism (SNP) chip의 SNP정보들을 활용한 친자 확인으로 혈통 정보의 신뢰도 향상에 기여하고자 실시하였다. 이미 혈통 정보가 확인되고 Microsatellite (MS) 유전자형으로친자 여부가 확인된 14개의 가계, 318두로 친자 확인 분석을 하였다. Call rate 100%인 마커들을 활용한 친자 확인 분석 결과, 9두가 모 부정,19두가 완전 부정으로 총 28두가 부정으로 판정되었고, 이는 부모의 SNP 정보에서 나올 수 있는 조합과 다른 자식의 유전자형이 확인되었다. 이후, 친자 확인 정확도 향상을 위해 call rate와 minor allele frequency (MAF)를 기준으로 SNP 마커 수를 증가시켜 분석하였으나 부정으로판정되는 개체들이 추가적으로 발생하였고, 이는 SNP 정보의 결측치 증가 및 자식의 유전자형 변이로 인한 것으로 판단된다. 따라서 고밀도SNP chip을 활용한 친자 확인 분석에는 보다 신중한 접근이 필요하며, 본 연구 결과는 고밀도 SNP 정보를 이용한 친자 확인 연구에 있어서기초자료로 활용될 수 있을 것으로 사료된다. This study was conducted to contribute to improving the reliability of pedigree information by confirming paternity test using SingleNucleotide Polymorphism (SNP) information from high-density SNP chips. Accurate pedigree information is becoming increasinglyimportant in the efforts to improve Hanwoo. As a result of a paternity test using markers with a 100% call rate, 318 animals wereanalyzed. A paternity test was conducted using 318 animals from 14 families whose pedigree information had already been confirmed,and their paternity was confirmed by the Microsatellite (MS) genotype. As a result of the paternity test using markers that are callrate of 100%, confirmed that the offspring's genotype is different from the combination of SNP information from the parents. Sincethen, the number of SNP markers has increased and has been analyzed based on call rate and minor allele frequency (MAF). However,it was found that additional individuals tested incorrect, which is believed to be due to an increase in missing SNP genotype informationand changes in the genotype of the offspring. Therefore, a more careful approach is needed for paternity testing using high-densitySNP chips, and the results of this study are believed to be used as foundational data for research on paternity testing using high-densitySNP information

전형매 가계 내 유전체정보량에 따른 유전체육종가 추정 및 정확도 분석

김은호,선두원,강호찬,명철현,김지영,이두호,이승환,임현태 경상국립대학교 농업생명과학연구원 2022 농업생명과학연구 Vol.56 No.6

The genomic estimated breeding value (GEBV) and accuracy of Korean beef (Hanwoo) are used as important indicators for selection, and recent studies using pedigree and genotype are being conducted to increase the reliability of the accuracy. Therefore, in this study, three families consisting of 10 Hanwoo with the same parents were collected to check the change in accuracy of individuals without genotypes according to the amount of genotypes in the family. GEBV and accuracy were estimated through single-step genomic best linear unbiased prediction (ssGBLUP) using 14,225 reference groups after assuming 5 test groups by selecting 10, 8, 6, 4, and 2 genotypes for each family. An H-matrix was constructed using pedigree and genotype of the groups, and the GEBV accuracy of carcass weight (CWT), eye muscle area (EMA), backfat thichness (BFT) and marbling score (MS) were estimated using the BLUPF90 program. Looking at the first family, the average GEBV accuracy of 10 heads with a genotype in the test group was estimated to be 0.734, 0.717, 0.712 and 0.745 in CWT, EMA, BFT and MS, respectively. Then, looking at the GEBV accuracy estimated by removing the genotype two by two, there was no change in accuracy in the case of individuals with a genotype, but the accuracy of the individuals without a genotype was estimated to be 0.114~0.168 lower on average. It was confirmed that the average GEBV accuracy of individuals without a genotype was 0.604~0.576, 0.6~0.573, 0.599~0.572 and 0.607~0.578 in CWT, EMA, BFT and MS, respectively. And it was confirmed that they decreased by 0.009 on average. Through this, although the amount of genotypes within a group did not have a significant effect on accuracy regardless of the presence of individual genotypes, the presence of individual genotypes had a greater effect on GEBV with high reliability. 한우 개량에 있어서 추정된 유전체육종가와 정확도는 선발에 중요한 지표로 사용되며, 최근 육종가 추정에 있어 정확도의 신뢰도를 높이기위해 혈통과 유전체정보를 이용한 연구가 활발히 진행되고 있다. 따라서, 본 연구는 가계 내 유전체정보량에 따라 유전체정보가 미포함된 개체의정확도 변화를 확인하고자 동일한 부모를 가진 한우 10두로 구성된 전형매 가계 3개를 수집하였으며, 각 가계 별로 유전체정보량을 10두, 8두, 6두, 4두, 2두씩 무작위로 선별하여 5가지의 검정집단으로 가정한 후 참조집단 14,225두를 이용하여 single step genomic best linear unbiased prediction (ssGBLUP)을 통해 genomic estimated breeding value (GEBV) 및 정확도를 추정하였다. 각 검정집단과 참조집단의 혈통 및 유전체정보를 이용하여 H-matrix를 구축하였고, BLUPF90 program을 사용하여 도체중, 등심단면적, 등지방두께, 근내지방도의 GEBV 및 정확도를 추정하였다. 첫 번째 가계를 대상으로 살펴보면, 검정집단에서 유전체정보를 보유하고 있는 10두의 GEBV 평균 정확도는 도체중 0.734, 등심단면적 0.717, 등지방두께 0.712, 근내지방도 0.745로 추정되었다. 이후 2두씩 무작위로 유전체정보를 제거하여 추정한 GEBV 정확도를 살펴보면, 유전체정보를보유한 개체의 경우 정확도의 변화가 나타나지 않았지만, 유전체정보가 미포함된 개체의 정확도가 평균 0.114 ~ 0.168 낮게 추정되었다. 가계내 유전체정보량에 따른 유전체정보가 미포함된 개체의 GEBV 평균 정확도는 도체중 0.604 ~ 0.576, 등심단면적 0.6 ~ 0.573, 등지방두께 0.599 ~ 0.572, 근내지방도 0.607 ~ 0.578로 평균 0.009씩 감소하는 것을 확인하였다. 이를 통해 가계 내 유전체정보량이 개체 별 유전체정보 유무와는상관없이 GEBV의 정확도 추정에 큰 영향이 없었으며, 신뢰도가 높은 GEBV 추정을 위해서는 개체 별 유전체정보의 유무가 더 큰 영향을 미친다는것을 확인하였다.