• Asian-Australasian Journal of Animal Sciences
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• 제33권2호
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• pp.203-211
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• 2020

Objective: Staphylococcus aureus (S. aureus) is one of the major microorganisms responsible for subclinical mastitis in dairy cattle. The present study was designed with the aim to explore the DNA methylation patterns using the Fluorescence-labeled methylation-sensitive amplified polymorphism (F-MSAP) techniques in a S. aureus-infected mouse model. Methods: A total of 12 out-bred Institute of Cancer Research female mice ranging from 12 to 13 weeks-old were selected to construct a mastitis model. F-MSAP analysis was carried out to detect fluctuations of DNA methylation between control group and S. aureus mastitis group. Results: Visible changes were observed in white cell counts in milk, percentage of granulocytes, percentage of lymphocytes, CD⁴⁺/CD⁸⁺ ratio (CD⁴⁺/CD⁸⁺), and histopathology of mice pre- and post-challenge with S. aureus. These findings showed the suitability of the S. aureus-infected mouse model. A total of 369 fragments was amplified from udder tissue samples from the two groups (S. aureus-infected mastitis group and control group) using eight pairs of selective primers. Results indicated that the methylation level of mastitis mouse group was higher than that in the control group. In addition, NCK-associated protein 5 (Nckap5) and transposon MTD were identified to be differentially methylated through secondary polymerase chain reaction and sequencing in the mastitis group. These observations might play an important role in the development of S. aureus mastitis. Conclusion: Collectively, our study suggests that the methylation modification in Nckap5 and transposon MTD might be considered as epigenetic markers in resistance to S. aureus-infected mastitis and provided a new insight into S. aureus mastitis research in dairy industry and public health.

• Asian-Australasian Journal of Animal Sciences
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• 제31권4호
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• pp.595-606
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• 2018

Objective: The Fusarium mycotoxins of deoxynivalenol (DON) and zerolenone (ZEN) cause health hazards for both humans and farm animals. Therefore, the main intention of this study was to reveal DON and ZEN effects on the mRNA expression of pro-inflammatory cytokines and other immune related genes in the liver of piglets. Methods: In the present study, 15 six-week-old piglets were randomly assigned to the following three different dietary treatments for 4 weeks: control diet, diet containing 8 mg DON/kg feed, and diet containing 0.8 mg ZEN/kg feed. After 4 weeks, liver samples were collected and sequenced using RNA-Seq to investigate the effects of the mycotoxins on genes and gene networks associated with the immune systems of the piglets. Results: Our analysis identified a total of 249 differentially expressed genes (DEGs), which included 99 upregulated and 150 downregulated genes in both the DON and ZEN dietary treatment groups. After biological pathway analysis, the DEGs were determined to be significantly enriched in gene ontology terms associated with many biological pathways, including immune response and cellular and metabolic processes. Consistent with inflammatory stimulation due to the mycotoxin-contaminated diet, the following Kyoto encyclopedia of genes and genomes pathways, which were related to disease and immune responses, were found to be enriched in the DEGs: allograft rejection pathway, cell adhesion molecules, graft-versus-host disease, autoimmune thyroid disease (AITD), type I diabetes mellitus, human T-cell leukemia lymphoma virus infection, and viral carcinogenesis. Genome-wide expression analysis revealed that DON and ZEN treatments downregulated the expression of the majority of the DEGs that were associated with inflammatory cytokines (interleukin 10 receptor, beta, chemokine [C-X-C motif] ligand 9), proliferation (insulin-like growth factor 1, major facilitator superfamily domain containing 2A, insulin-like growth factor binding protein 2, lipase G, and salt inducible kinase 1), and other immune response networks (paired immunoglobulin-like type 2 receptor beta, Src-like-adaptor-1 [SLA1], SLA3, SLA5, SLA7, claudin 4, nicotinamide N-methyltransferase, thyrotropin-releasing hormone degrading enzyme, ubiquitin D, histone $H_2B$ type 1, and serum amyloid A). Conclusion: In summary, our results demonstrated that high concentrations DON and ZEN disrupt immune-related processes in the liver.

• Asian-Australasian Journal of Animal Sciences
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• 제30권11호
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• pp.1529-1539
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• 2017

Objective: The objective of this study was to compare the DNA methylation profile in the longissimus dorsi muscle between Small Tailed Han and Dorper${\times}$Small Tailed Han crossbred sheep which were known to exhibit significant difference in meat-production. Methods: Six samples (three in each group) were subjected to the methylated DNA immunoprecipitation sequencing (MeDIP-seq) and subsequent bioinformatics analyses to detect differentially methylated regions (DMRs) between the two groups. Results: 23.08 Gb clean data from six samples were generated and 808 DMRs were identified in gene body or their neighboring up/downstream regions. Compared with Small Tailed Han sheep, we observed a tendency toward a global loss of DNA methylation in these DMRs in the crossbred group. Gene ontology enrichment analysis found several gene sets which were hypomethylated in gene-body region, including nucleoside binding, motor activity, phospholipid binding and cell junction. Numerous genes were found to be differentially methylated between the two groups with several genes significantly differentially methylated, including transforming growth factor beta 3 (TGFB3), acyl-CoA synthetase long chain family member 1 (ACSL1), ryanodine receptor 1 (RYR1), acyl-CoA oxidase 2 (ACOX2), peroxisome proliferator activated receptor-gamma2 (PPARG2), netrin 1 (NTN1), ras and rab interactor 2 (RIN2), microtubule associated protein RP/EB family member 1 (MAPRE1), ADAM metallopeptidase with thrombospondin type 1 motif 2 (ADAMTS2), myomesin 1 (MYOM1), zinc finger, DHHC type containing 13 (ZDHHC13), and SH3 and PX domains 2B (SH3PXD2B). The real-time quantitative polymerase chain reaction validation showed that the 12 genes are differentially expressed between the two groups. Conclusion: In the current study, a tendency to a global loss of DNA methylation in these DMRs in the crossbred group was found. Twelve genes, TGFB3, ACSL1, RYR1, ACOX2, PPARG2, NTN1, RIN2, MAPRE1, ADAMTS2, MYOM1, ZDHHC13, and SH3PXD2B, were found to be differentially methylated between the two groups by gene ontology enrichment analysis. There are differences in the expression of 12 genes, of which ACSL1, RIN2, and ADAMTS2 have a negative correlation with methylation levels and the data suggest that DNA methylation levels in DMRs of the 3 genes may have an influence on the expression. These results will serve as a valuable resource for DNA methylation investigations on screening candidate genes which might be related to meat production in sheep.

• Asian-Australasian Journal of Animal Sciences
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• 제31권1호
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• pp.138-148
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• 2018

Objective: Fusarium mycotoxins deoxynivalenol (DON) and zearalenone (ZEN), common contaminants in the feed of farm animals, cause immune function impairment and organ inflammation. Consequently, the main objective of this study was to elucidate DON and ZEN effects on the mRNA expression of pro-inflammatory cytokines and other immune related genes in the kidneys of piglets. Methods: Fifteen 6-week-old piglets were randomly assigned to three dietary treatments for 4 weeks: control diet, and diets contaminated with either 8 mg DON/kg feed or 0.8 mg ZEN/kg feed. Kidney samples were collected after treatment, and RNA-seq was used to investigate the effects on immune-related genes and gene networks. Results: A total of 186 differentially expressed genes (DEGs) were screened (120 upregulated and 66 downregulated). Gene ontology analysis revealed that the immune response, and cellular and metabolic processes were significantly controlled by these DEGs. The inflammatory stimulation might be an effect of the following enriched Kyoto encyclopedia of genes and genomes pathway analysis found related to immune and disease responses: cytokine-cytokine receptor interaction, chemokine signaling pathway, toll-like receptor signaling pathway, systemic lupus erythematosus (SLE), tuberculosis, Epstein-Barr virus infection, and chemical carcinogenesis. The effects of DON and ZEN on genome-wide expression were assessed, and it was found that the DEGs associated with inflammatory cytokines (interleukin 10 receptor, beta, chemokine [C-X-C motif] ligand 9, CXCL10, chemokine [C-C motif] ligand 4), proliferation (insulin like growth factor binding protein 4, IgG heavy chain, receptor-type tyrosine-protein phosphatase C, cytochrome P450 1A1, ATP-binding cassette sub-family 8), and other immune response networks (lysozyme, complement component 4 binding protein alpha, oligoadenylate synthetase 2, signaling lymphocytic activation molecule-9, ${\alpha}$-aminoadipic semialdehyde dehydrogenase, Ig lambda chain c region, pyruvate dehydrogenase kinase, isozyme 4, carboxylesterase 1), were suppressed by DON and ZEN. Conclusion: In summary, our results indicate that high concentrations of DON and ZEN suppress the inflammatory response in kidneys, leading to potential effects on immune homeostasis.

• Journal of the Korean Data and Information Science Society
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• 제25권5호
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• pp.1057-1067
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• 2014

인간게놈 프로젝트 이후 질병과 연관된 변이유전자를 탐색하기 위해 유전형질의 차이에 영향을 주는 단일 유전자를 중심으로 전장유전체 연관성 연구가 활발하게 진행되어왔다. 그러나 전장유전체 연관성 연구에서 접근한 단일유전자 분석방법에 한계점이 발견되면서 최근에는 다중유전자 분석방법이나 유전자-유전자간의 상호작용에 대한 연구들이 활발하게 진행 중이다. 이 중 다중차원축소방법은 유전자-유전자간의 상호작용의 연관성을 찾아내기 위하여 고차원을 일차원으로 축소하는 방법으로 이진형 반응변수를 기반으로 크게 활용되고 있다. 본 논문에서는 이 방법을 생존시간으로 확장하여 생존시간과 연관된 유전자-유전자간의 상호작용을 찾아내는 방법을 제안하고자 한다. 구체적으로 가속화 고장시간 회귀모형 하에서 표준화잔차 스코어를 분류기준으로 사용하여 다중차원축소방법을 적용하는 방법으로 AFT-MDR이라고 지칭하였다. 시뮬레이션 연구를 통하여 기존에 제안된 Surv-MDR과 Cox-MDR과의 검정력을 비교하였으며 국내의 백혈병환자 자료분석에 적용하였다. 시뮬레이션 결과로부터 AFT-MDR은 유전율이 높을수록 검정력이 커지며 회귀모형에 기반하여 공변량의 효과를 고려할 수 있다는 장점이 있으나 센서링 비율이 높아지면서 검정력이 매우 떨어진다는 단점이 발견되었다. 따라서 이를 보완하기 위한 추후연구의 필요성이 요구된다.

• Animal Bioscience
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• 제34권12호
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• pp.1879-1885
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• 2021

Objective: Owing to the lack of a breeding index for efficient and quick fertility evaluations of Holstein bulls when using traditional or genome-wide detection methods, this study aimed to determine whether in vitro fertilization (IVF) could be used as an indicator of conception rate of artificial insemination (AI). Methods: Conventional and sexed frozen semen from nine bulls were used for IVF and AI. Results: The IVF and AI conception rates of each bull were confirmed to be positively correlated between the conventional frozen and sexed frozen semen. The correlation coefficient R values of nine bulls between IVF and AI methods were 0.73 and 0.97 for the conventional frozen and sexed frozen semen, respectively. The average conception rate of three bulls undergoing AI was 69.5% and 64.2%, 61.8% and 58.8%, and 48.2% and 46.2% in first-, second-, and third-born cows when conventional frozen and sexed frozen semen were used, respectively, which showed a positive correlation with the fertilization rate in the same parity. We propose an evaluation standard to assess the fertilization ability of bulls based on their IVF test results, which is categorized into three grades: grade one, normal fertility bull with an AI conception rate of 40%±5% and IVF rate of 45% to 60%; grade two, higher fertility bull with an AI conception rate of 50%±5% and IVF rate of 61% to 80%; and grade three, highest fertility bull with an AI conception rate of 60%±5% and IVF rate of >80%. Conclusion: These findings reveal that IVF results can be used as a breeding index for bulls to evaluate their AI conception ability, which may shorten the time required to select bulls for breeding.

• Animal Bioscience
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• 제34권11호
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• pp.1739-1748
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• 2021

Objective: In recent years, long noncoding RNAs (lncRNAs) have been identified in many species, and some of them have been shown to play important roles in muscle development and myogenesis. However, the differences in lncRNAs between Kazakh cattle and Xinjiang brown cattle remain undefined; therefore, we aimed to confirm whether lncRNAs are differentially expressed in the longissimus dorsi between these two types of cattle and whether differentially expressed lncRNAs regulate muscle differentiation. Methods: We used RNA-seq technology to identify lncRNAs in longissimus muscles from these cattle. The expression of lncRNAs were analyzed using StringTie (1.3.1) in terms of the fragments per kilobase of transcript per million mapped reads values of the encoding genes. The differential expression of the transcripts in the two samples were analyzed using the DESeq R software package. The resulting false discovery rate was controlled by the Benjamini and Hochberg's approach. KOBAS software was utilized to measure the expression of different genes in Kyoto encyclopedia of genes and genomes pathways. We randomly selected eight lncRNA genes and validated them by quantitative reverse transcription polymerase chain reaction (RT-qPCR). Results: We found that 182 lncRNA transcripts, including 102 upregulated and 80 downregulated transcripts, were differentially expressed between Kazakh cattle and Xinjiang brown cattle. The results of RT-qPCR were consistent with the sequencing results. Enrichment analysis and functional annotation of the target genes revealed that the differentially expressed lncRNAs were associated with the mitogen-activated protein kinase, Ras, and phosphatidylinositol 3-kinase (PI3k)/Akt signaling pathways. We also constructed a lncRNA/mRNA coexpression network for the PI3k/Akt signaling pathway. Conclusion: Our study provides insights into cattle muscle-associated lncRNAs and will contribute to a more thorough understanding of the molecular mechanism underlying muscle growth and development in cattle.

• 한국산림과학회지
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• 제112권1호
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• pp.40-56
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• 2023

사시나무속 수종은 생장이 빠르고 우수한 탄소흡수 능력을 보여주며, 환경정화 효과가 큰 수종으로 이상기후 및 환경오염 문제에 대응하는 기후적응성 품종개발 및 육종집단 조성에 적합하다. 따라서 유전연관지도 작성 및 양적형질 유전자좌 탐색을 통하여 포플러 육종을 신속하게 진행할 수 있을 것이다. 본 연구에서는 차세대 염기서열 분석기술 방법인 genotyping-by-sequencing 기법을 이용해 인공교배 차대에 대한 고밀도 유전연관 지도를 작성하였다. 또한 사시나무의 수고와 근원경 생장 그리고 해충피해에 대한 회복력 형질을 조사하여 유전연관지도에 위치한 양적형질 유전자좌를 탐색하였다. 서울대학교 학술림에 조성된 사시나무 4년생 육종집단(오대19 × 봉현4 인공교배 차대집단)에서 수고 및 근원경 생장을 조사하였으며, 식엽성 해충인 꼬마버들재주나방 유충의 피해를 받은 후 이에 대해 회복 능력을 조사하였다. 잎 시료의 DNA 추출 후 5개 microsatellite 마커를 이용하여 유전자형을 확인하였으며 친자로 확인된 개체만을 연구재료로 사용하였다. 친자 확인이 완료된 시료의 DNA는 제한효소를 이용해 절단하였으며, 이렇게 얻은 DNA 조각들은 GBS 라이브러리로 제작하여 염기서열을 분석하였다. 분석된 결과는 Populus trichocarpa를 참조유전체로 하여 정렬하였다. 정렬된 SNP 마커는 총 58,040개였으며, 그 가운데 17,755개의 SNP 마커를 유전연관지도 작성에 사용하였다. 유전연관지도는 19개의 연관군으로 나누어졌으며, 전체 길이는 2,129.54 cM으로 나타났다. 조사된 세 가지 형질에 대한 양적형질 유전자좌 분석을 실시한 결과, 수고와 근원경 생장과 연관된 양적형질 유전자좌는 찾을 수 없었으나 전장유전체연관연구(GWAS)를 통하여 4번 연관군(염색체)에 해충피해 회복력과 관련이 있을 것으로 추정되는 유전자를 확인하였다.

• Asian Pacific Journal of Cancer Prevention
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• 제13권12호
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• pp.6273-6276
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• 2012

Background/Aim: Six prostate cancer (PCa) susceptibility loci were identified in a genome-wide association study (GWAS) in populations of European decent. However, the associations of these 6 single-nucleotide polymorphisms (SNPs) with PCa has remained tobe clarified in men in Northern China. This study aimed to explore the loci associated with PCa risk in a Northern Chinese population. Methods: Blood samples and clinical information of 289 PCa patients and 288 controls from Beijing and Tianjin were collected. All risk SNPs were genotyped using polymerase chain reaction (PCR)-high resolution melting curve technology and gene sequencing. Associations between PCa and clinical covariates (age at diagnosis, prostate-specific antigen [PSA], Gleason score, tumor stage, and level of aggressiveness) and frequencies of alleles and genotypes of these SNPs were analyzed using genetic statistics. Results: Among the candidate SNPs, 11p15 (rs7127900, A) was associated with PCa risk (P = 0.02, odds ratio [OR] = 1.64, 95% confidence interval [CI] = 1.09-2.46). Genotypes showed differences between cases and controls on 11p15 (rs7127900, A), 11q13 (rs7931342, T), and HNF1B (rs4430796, A) (P = 0.03, P = 0.01, and P = 0.04, respectively). The genotype TG on 11q13 (rs7931342, T) was positively associated with an increased Gleason score (P = 0.04, OR = 2.15, 95% CI = 1.02-4.55). Patients carrying TG on 17q24 (rs1859962, G) were negatively associated with an increased body mass index (BMI) (P = 0.03, OR = 0.44, 95% CI = 0.21-0.92) while those with AG on HNF1B (rs4430796, A) were more likely to have PSA increase (P = 0.002). Conclusion: Our study suggests that 11p15 (rs7127900, A) could be a susceptibility locus associated with PCa in Northern Chinese. Genotype TG on 11q13 (rs7931342, T) could be related to an increased Gleason score, AG on HNF1B (rs4430796, A) could be associated with PSA increase, and TG on 17q24 (rs1859962, G) could be negatively associated with an increased BMI in Chinese men with PCa.