Genome-wide association study identifies genomic regions associated with key reproductive traits in Korean Hanwoo cows.

BACKGROUND: Conducting genome-wide association studies (GWAS) for reproductive traits in Hanwoo cattle, including age at first calving (AFC), calving interval (CI), gestation length (GL), and number of artificial inseminations per conception (NAIPC), is of paramount significance. These analyses provided a thorough exploration of the genetic basis of these traits, facilitating the identification of key markers for targeted trait improvement. Breeders can optimize their selection strategies, leading to more efficient and sustainable breeding programs, by incorporating genetic insights. This impact extends beyond individual traits and contributes to the overall productivity and profitability of the Hanwoo beef cattle industry. Ultimately, GWAS is essential in ensuring the long-term genetic resilience and adaptability of Hanwoo cattle populations. The primary goal of this study was to identify significant single nucleotide polymorphisms (SNPs) or quantitative trait loci (QTLs) associated with the studied reproductive traits and subsequently map the underlying genes that hold promise for trait improvement. RESULTS: A genome-wide association study of reproductive traits identified 68 significant single nucleotide polymorphisms (SNPs) distributed across 29 Bos taurus autosomes (BTA). Among them, BTA14 exhibited the highest number of identified SNPs (25), whereas BTA6, BTA7, BTA8, BTA10, BTA13, BTA17, and BTA20 exhibited 8, 5, 5, 3, 8, 2, and 12 significant SNPs, respectively. Annotation of candidate genes within a 500 kb region surrounding the significant SNPs led to the identification of ten candidate genes relevant to age at first calving. These genes were: FANCG, UNC13B, TESK1, TLN1, and CREB3 on BTA8; FAM110B, UBXN2B, SDCBP, and TOX on BTA14; and MAP3K1 on BTA20. Additionally, APBA3, TCF12, and ZFR2, located on BTA7 and BTA10, were associated with the calving interval; PAX1, SGCD, and HAND1, located on BTA7 and BTA13, were linked to gestation length; and RBM47, UBE2K, and GPX8, located on BTA6 and BTA20, were linked to the number of artificial inseminations per conception in Hanwoo cows. CONCLUSIONS: The findings of this study enhance our knowledge of the genetic factors that influence reproductive traits in Hanwoo cattle populations and provide a foundation for future breeding strategies focused on improving desirable traits in beef cattle. This research offers new evidence and insights into the genetic variants and genome regions associated with reproductive traits and contributes valuable information to guide future efforts in cattle breeding.

Rearing characteristics of fattening Hanwoo steers managed in different stocking densities.

OBJECTIVE: This study was conducted to analyze the effects of stocking density on growth and carcass quality, and behavior of Hanwoo cattle to conform with global trends, targeting animal welfare production through the practice of environmentally friendly condition. METHODS: Thirty six steers were randomly assigned to three treatment groups (C: 5 heads, T1: 4 heads, T2: 3 heads) and reared in separate pens with a constant stocking density of 50 m2 (C: 10 m2/head, T1: 12.5 m2/head, T2: 16.67 m2/head) per group from 12 to 30 month of age. Growth performance, behavior and carcass quality traits of each steer were recorded and compared between the treatment groups. RESULTS: In general, the average daily gain during the fattening period was lower in group T2 than in T1 and the control groups. However, carcass weight and dressing percentage was lower in the control group than in T1 or T2 groups (p<0.05). Also, marbling score at 30 months of age was the lowest in the control group (p<0.05), while the three heads group (T2) had the greatest longissimus muscle area and marbling score (p<0.05). The behavior of walking time was the greatest in T2 group, while self-grooming and fighting occurred with the most frequency in the control group (p<0.05). CONCLUSION: Our results show that the steers in more spacious stocking density had better carcass quality and wellbeing related behaviors, indicating that a lower density has a positive effect on raising management and carcass quality. Thus it is a need to install appropriate pens fitted to welfare-oriented management practices from growing to fattening period in Hanwoo cattle.

Relationship between Single Nucleotide Polymorphisms in the Peroxisome Proliferator-Activated Receptor Gamma Gene and Fatty Acid Composition in Korean Native Cattle.

The peroxisome proliferator-activated receptor gamma (PPARγ) gene plays an important role in the biosynthesis process controlled by a number of fatty acid transcription factors. This study investigates the relationships between 130 single-nucleotide polymorphisms (SNPs) in the PPARγ gene and the fatty acid composition of muscle fat in the commercial population of Korean native cattle. We identified 38 SNPs and verified relationships between 3 SNPs (g.1159-71208 A>G, g.42555-29812 G>A, and g.72362 G>T) and the fatty acid composition of commercial Korean native cattle (n = 513). Cattle with the AA genotype of g.1159-71208 A>G and the GG genotype of g.42555-29812 G>A and g.72362 G>T had higher levels of monounsaturated fatty acids and carcass traits (p<0.05). The results revealed that the 3 identified SNPs in the PPARγ gene affected fatty acid composition and carcass traits, suggesting that these 3 SNPs may improve the flavor and quality of beef in commercial Korean native cattle.

Identification of exonic nucleotide variants of the thyroid hormone responsive protein gene associated with carcass traits and Fatty Acid composition in korean cattle.

The thyroid hormone responsive protein (THRSP) gene is a functional gene that can be used to indicate the fatty acid compositions. This study investigates the relationships of exonic single nucleotide polymorphisms (SNPs) in the THRSP gene and fatty acid composition of muscle fat and marbling score in the 612 Korean cattle. The relationships between fatty acid composition and eight SNPs in the THRSP gene (g.78 G>A, g.173 C>T, g.184 C>T, g.190 C>A, g.194 C>T, g.277 C>G, g.283 T>G and g.290 T>G) were investigated, and according to the results, two SNPs (g.78 G>A and g.184 C>T) in exon 1 were associated with fatty acid composition. The GG and CC genotypes of g.78 G>A and g.184 C>T had higher unsaturated fatty acid (UFA) and monounsaturated fatty acid (MUFA) content (p<0.05). In addition, the ht1*ht1 group (Val/Ala haplotype) in a linkage disequilibrium increased MUFAs and marbling scores for carcass traits (p<0.05). As a result, g.78 G>A and g.184 C>T had significantly relationships with UFAs and MUFAs. Two SNPs in the THRSP gene affected fatty acid composition, suggesting that GG and CC genotypes and the ht1*ht1 group (Val/Ala haplotype) can be markers to genetically improve the quality and flavor of beef.

Application of sperm motion kinematics and motility-related proteins for prediction of male fertility.

The selection of superior sires is paramount for enhancing the efficiency of animal production in the livestock industry. However, semen quality assessment still relies on conventional semen analysis techniques in both animals and humans. Despite extensive efforts to develop various biomarkers for more accurate and precise predictions of male fertility potential, more effective physiological indicators and advance potential biomarkers are needed. Herein, we aimed to develop new potential biomarkers related to sperm motion kinematics for male fertility prediction. We first evaluated sperm motion kinematic parameters and expression levels of sperm motility-related proteins of 30 Duroc boars. We then explored the correlation between litter size, sperm motion kinematics parameters, and sperm motility-related proteins. Progressive sperm motility (%), rapid sperm motility (%), slow sperm motility (%), straight-line velocity (µm/s), linearity (%), beat cross frequency (Hz), mean angular displacement (degree), wobble (%) were correlated with litter size. Furthermore, the expression of axonemal dynein light intermediate polypeptide 1 (DNALI1) and radial spoke head protein 9 homolog (RSPH9) correlated with litter size. The overall accuracy exceeded 60% for predicting litter size using these sperm motion parameters and proteins. Notably, our study observed an increase in litter size after predicting litter size using these parameters and proteins. Thus, sperm motion kinematic parameters and protein expression, particularly of DNALI1 and RSPH9, could serve as new biomarkers for male fertility. These results may contribute to improved understanding of the mechanisms underlying sperm motility.

Adverse Effects of Avobenzone on Boar Sperm Function: Disruption of Protein Kinase A Activity and Tyrosine Phosphorylation.

Avobenzone (AVO), an ultraviolet (UV) filter, is frequently used as an ingredient in personal cosmetics. This UV filter has been found to be easily exposed in swimming pools and beaches, and it has been detected in human urine and blood. Moreover, numerous studies have demonstrated that AVO exhibits endocrine-disrupting properties. Nevertheless, the effects of AVO on male fertility have not yet fully understood. Therefore, this study aimed to assess the effects of AVO on various sperm functions during capacitation. First, boar spermatozoa were treated with various AVO concentrations. After treatment, sperm motility and kinetic characteristics, capacitation status, intracellular adenosine triphosphate (ATP) levels, and sperm viability were evaluated. Moreover, Western blot analysis w.as conducted to evaluate protein kinase A (PKA) activity and tyrosine phosphorylation. As a result, AVO treatment significantly decreased total motility, progressive motility, and several kinetic characteristics at high concentrations (50 and 100 µM). Furthermore, the capacitation status dose-dependently decreased. Conversely, no significant differences in acrosome reaction, cell viability, and intracellular ATP levels were observed. However, the intracellular ATP level tended to decrease. In addition, AVO dose-dependently induced abnormal changes in PKA activity and tyrosine phosphorylation. Although AVO did not directly exert a toxic effect on cell viability, it ultimately negatively affected sperm functions through abnormal alterations in PKA activity and tyrosine phosphorylation. Thus, the potential implications on male fertility must be considered when contemplating the safe utilization of AVO.

Effects of Oridonin on Sperm Function and the PI3K/PDK1/AKT Signaling Pathway: Implications for Reproductive Toxicity.

Oridonin, a natural terpenoid isolated from the leaves of Isodon rubescens (Hemsley) H.Hara, is widely used in oriental medicine for its anticancer properties across various cancer types. Despite its prevalent use, the toxic effects of oridonin on male reproduction, particularly its impact on sperm functions and the mechanisms involved, are not well understood. This study aimed to explore the effects and underlying mechanisms of oridonin on sperm functions. We initially treated Duroc boar spermatozoa with varying concentrations of oridonin (0, 5, 50, 75, 100, and 150µM) and incubated them to induce capacitation. We then assessed cell viability and several sperm functions, including sperm motility and motion kinematics, capacitation status, and ATP levels. We also analyzed the expression levels of proteins associated with the phosphatidylinositol 3-kinase (PI3K)/phosphoinositide-dependent kinase-1 (PDK1)/protein kinase B (AKT) signaling pathway and phosphotyrosine proteins. Our results indicate that oridonin adversely affects most sperm functions in a dose-dependent manner. We observed significant decreases in AKT, p-AKT (Thr308), phosphatase and tensin homolog (PTEN), p-PDK1, and p-PI3K levels following oridonin treatment, alongside an abnormal increase in phosphotyrosine proteins. These findings suggest that oridonin may disrupt normal levels of tyrosine-phosphorylated proteins by inhibiting the PI3K/PDK1/AKT signaling pathway, which is crucial for cell proliferation, metabolism, and apoptosis, thus potentially harming sperm functions. Consequently, we recommend considering the reproductive toxicity of oridonin when using it as a therapeutic agent.

Identification of Stearoyl-CoA Desaturase (SCD) Gene Interactions in Korean Native Cattle Based on the Multifactor-dimensionality Reduction Method.

Fat quality is determined by the composition of fatty acids. Genetic relationships between this composition and single nucleotide polymorphisms (SNPs) in the stearoyl-CoA desaturase1 (SCD1) gene were examined using 513 Korean native cattle. Single and epistatic effects of 7 SNP genetic variations were investigated, and the multifactor dimensionality reduction (MDR) method was used to investigate gene interactions in terms of oleic acid (C18:1), mono-unsaturated fatty acids (MUFAs) and marbling score (MS). The g.6850+77 A>G and g.14047 C>T SNP interactions were identified as the statistically optimal combination (C18:1, MUFAs and MS permutation p-values were 0.000, 0.000 and 0.001 respectively) of two-way gene interactions. The interaction effects of g.6850+77 A>G, g.10213 T>C and g.14047 C>T reflected the highest training-balanced accuracy (63.76%, 64.70% and 61.85% respectively) and was better than the individual effects for C18:1, MUFAs and MS. In addition, the superior genotype groups were AATTCC, AGTTCC, GGTCCC, AGTCCT, GGCCCT and AGCCTT. These results suggest that the selected SNP combination of the SCD1 gene and superior genotype groups can provide useful inferences for the improvement of the fatty acid composition in Korean native cattle.

Genomic Predictions in Korean Hanwoo Cows: A Comparative Analysis of Genomic BLUP and Bayesian Methods for Reproductive Traits.

This study aimed to predict the accuracy of genomic estimated breeding values (GEBVs) for reproductive traits in Hanwoo cows using the GBLUP, BayesB, BayesLASSO, and BayesR methods. Accuracy estimates of GEBVs for reproductive traits were derived through fivefold cross-validation, analyzing a dataset comprising 11,348 animals and employing an Illumina Bovine 50K SNP chip. GBLUP showed an accuracy of 0.26 for AFC, while BayesB, BayesLASSO, and BayesR demonstrated values of 0.28, 0.29, and 0.29, respectively. For CI, GBLUP attained an accuracy of 0.19, whereas BayesB, BayesLASSO, and BayesR scored 0.21, 0.24, and 0.25, respectively. The accuracy for GL was uniform across GBLUP, BayesB, and BayesR at 0.31, whereas BayesLASSO showed a slightly higher accuracy of 0.33. For NAIPC, GBLUP showed an accuracy of 0.24, while BayesB, BayesLASSO, and BayesR recorded 0.22, 0.27, and 0.30, respectively. The variation in genomic prediction accuracy among methods indicated Bayesian approaches slightly outperformed GBLUP. The findings suggest that Bayesian methods, notably BayesLASSO and BayesR, offer improved predictive capabilities for reproductive traits. Future research may explore more advanced genomic approaches to enhance predictive accuracy and genetic gains in Hanwoo cattle breeding programs.

The natural flavonoid compound deguelin suppresses sperm (Sus Scrofa) functions through abnormal activation of the PI3K/AKT pathway.

Deguelin is a natural flavonoid extracted from plants belonging to the Lonchocarpus, Derris, or Tephrosia genera. It inhibits AKT activity in tumors and has the potential to be used as a treatment for malignant tumors. However, the risks associated with the use of deguelin on male fertility have not yet been explained in detail. Therefore, this study was conducted to investigate the effects of deguelin on sperm functions during capacitation. First, boar spermatozoa were exposed to different concentrations of deguelin (0.1, 1, 10, 50, and 100 µM). Next, sperm functional assessments, such as sperm motility, capacitation status, intracellular ATP level, and cell viability, were performed. The expression levels of PI3K/AKT-related proteins and the phosphorylation of their tyrosine residues were also evaluated by western blotting. No significant difference was observed in cell viability; however, deguelin considerably decreased sperm motility and motion kinematics in a dose-dependent manner. Although no significant difference was observed in the capacitation status, acrosome reaction decreased at high concentrations of deguelin (50 and 100 µM). Furthermore, intracellular ATP levels were significantly decreased in all deguelin treatment groups compared with those in the control group. Results of western blotting revealed that deguelin substantially diminished tyrosine phosphorylation. Interestingly, in contrast to previous studies showing that deguelin inhibits AKT activity, our results showed that it increased the expression of PI3K/AKT pathway-related proteins. Collectively, these findings indicate that deguelin exerts negative effects on sperm functions due to abnormal PI3K/AKT signaling activation. We believe that this is the first study to provide evidence that deguelin can regulate sperm functions independent of PI3K/AKT pathway inhibition. Furthermore, its detrimental effects on male fertility should be considered while developing or using deguelin as a therapeutic agent.

Genome-Wide Association Study to Identify QTL for Carcass Traits in Korean Hanwoo Cattle.

This study aimed to identify genetic associations with carcass traits in Hanwoo cattle using a genome-wide association study. A total of 9302 phenotypes were analyzed, and all animals were genotyped using the Illumina Bovine 50K v.3 SNP chip. Heritabilities for carcass weight (CWT), eye muscle area (EMA), backfat thickness (BF), and marbling score (MS) were estimated as 0.42, 0.36, 0.36, and 0.47, respectively, using the GBLUP model, and 0.47, 0.37, 0.36, and 0.42, respectively, using the Bayes B model. We identified 129 common SNPs using DGEBV and 118 common SNPs using GEBV on BTA6, BTA13, and BTA14, suggesting their potential association with the traits of interest. No common SNPs were found between the GBLUP and Bayes B methods when using residuals as a response variable in GWAS. The most promising candidate genes for CWT included SLIT2, PACRGL, KCNIP4, RP1, XKR4, LYN, RPS20, MOS, FAM110B, UBXN2B, CYP7A1, SDCBP, NSMAF, TOX, CA8, LAP3, FAM184B, and NCAPG. For EMA, the genes IBSP, LAP3, FAM184B, LCORL, NCAPG, SLC30A9, and BEND4 demonstrated significance. Similarly, CYP7B1, ARMC1, PDE7A, and CRH were associated with BF, while CTSZ, GNAS, VAPB, and RAB22A were associated with MS. This finding offers valuable insights into genomic regions and molecular mechanisms influencing Hanwoo carcass traits, aiding efficient breeding strategies.

Generation of double knockout cattle via CRISPR-Cas9 ribonucleoprotein (RNP) electroporation.

BACKGROUND: Genome editing has been considered as powerful tool in agricultural fields. However, genome editing progress in cattle has not been fast as in other mammal species, for some disadvantages including long gestational periods, single pregnancy, and high raising cost. Furthermore, technically demanding methods such as microinjection and somatic cell nuclear transfer (SCNT) are needed for gene editing in cattle. In this point of view, electroporation in embryos has been risen as an alternative. RESULTS: First, editing efficiency of our electroporation methods were tested for embryos. Presence of mutation on embryo was confirmed by T7E1 assay. With first combination, mutation rates for MSTN and PRNP were 57.6% ± 13.7% and 54.6% ± 13.5%, respectively. In case of MSTN/BLG, mutation rates were 83.9% ± 23.6% for MSTN, 84.5% ± 18.0% for BLG. Afterwards, the double-KO embryos were transferred to surrogates and mutation rate was identified in resultant calves by targeted deep sequencing. Thirteen recipients were transferred for MSTN/PRNP, 4 calves were delivered, and one calf underwent an induction for double KO. Ten surrogates were given double-KO embryos for MSTN/BLG, and four of the six calves that were born had mutations in both genes. CONCLUSIONS: These data demonstrated that production of genome edited cattle via electroporation of RNP could be effectively applied. Finally, MSTN and PRNP from beef cattle and MSTN and BLG from dairy cattle have been born and they will be valuable resources for future precision breeding.

Behavioral changes of sows with changes in flattening rate.

In this study, considering the difficulties for all farms to convert farm styles to animal welfare-based housing, an experiment was performed to observe the changes in the behavior and welfare of sows when the slat floor was changed to a collective breeding ground. Twenty-eight sows used in this study were between the second and fifth parities to minimize the influence of parity. Using a flats floor cover, the flattening rates were treated as 0%, 20%, 30%, 40%, and 50%. Data collection was the behavior of sows visually observed using a camera (e.g., standing, lying, fighting and excessive biting behaviors, and abnormal behaviors) and the animal welfare level measured through field visits. Lying behavior was found to be higher (p < 0.01) as the flattening rate increased, and sows lying on the slatted cover also increased as the flattening rate increased (p < 0.01). Fighting behavior wasincreased when the flattening rate was increased to 20%, and chewing behavior was increased (p < 0.05) as the flattening rate increased. The animal welfare level of sows, 'good feeding', it was found that all treatment groups for body condition score and water were good at 100 (p < 0.05). 'Good housing' was the maximum value (100) in each treatment group. As the percentage of floor increased, the minimum good housing was increased from 78 in 0% flattening rate to 96 in 50% flattening rate. The maximum (100) 'good health' was achieved in the 0% and 20% flattening rates, and it was 98, 98, and 99 in the 30%, 50%, and 40% flattening rate, respectively. 'Appropriate behavior' score was significantly lower than that of other paremeters, but when the flattening ratio was 0% and 20%, the maximum and minimum values were 10. At 40% and 50%, the maximum values were 39 and 49, respectively, and the minimum values were analyzed as 19 for both 40% and 50%. These results will be used as basic data about sow welfare for farmers to successfully transition to group housing and flat floors.

Effect of glycerol addition time on the cryopreserved Korean native brindle cattle (Chikso) sperm quality.

Although cryopreservation is an efficient method for maintaining the biological and genetic resources of sperm, the sperm damage during the cryopreservation process cannot be ignored. It should be possible to obtain the most effective cryopreservation performance by accurately grasping the effects of various factors on the cryopreservation of sperm. The previous study demonstrated that a suitable standard protocol for cryopreservation of Korean native brindled cattle (Chikso) does not exist, based on the methods for semen cryopreservation of Chikso differ in each research center. The most obvious difference between most of protocols is the addition of glycerol before and after cooling during the Chikso cryopreserved semen process. Therefore we focused on the effects of glycerol addition time on the quality of cryopreserved Chikso sperm. In the present study, 27 individual Chikso samples were collected by transrectal massage and divided into two parts: the "cryopreservation method A" group (adding glycerol before cooling) and the "cryopreservation method B" group (adding glycerol after cooling). Meanwhile, the values of various sperm parameters were derived from each group, including sperm motility, kinematics, capacitation status, cell viability, and intracellular ATP levels, which we used to compare and evaluate sperm function. The results of this study indicated that during the semen cryopreservation process of the Chikso, the addition of glycerol after cooling yielded superior results in a variety of sperm parameters, such as sperm motility, progressive motility, rapid motility, VCL, VSL, VAP, ALH, capacitation status, viability, and intracellular ATP level after freezing and thawing. Our study is suggested that the glycerol addition time during the cryopreservation process for Chikso should be considered. In addition, our results may be provided reference to develop suitable the cryopreservation procedure of the Chikso sperm.

Ras-related proteins (Rab) play significant roles in sperm motility and capacitation status.

Rab proteins are widely known for their involvement in establishing Golgi apparatus and controlling Golgi trafficking in eukaryotic cells. Specifically, Rab proteins play significant roles in acrosome formation and exocytosis. Furthermore, mechanisms involved in the regulation of Rab proteins during capacitation have been identified. However, there has been no direct evaluation to assess the correlation between Rab proteins and sperm function. Consequently, this study was designed to analyze the correlation between Rab proteins and sperm functions. Individually, we analyzed the sperm motility patterns, motion kinematics, capacitation status, and Rab protein expression levels of sperm samples from 31 boars before and after capacitation. As a result, we discovered that Rab3A, Rab5, Rab11, Rab14, and Rab27A correlated with various sperm motility patterns, motion kinematics before capacitation. Rab3A, Rab5, Rab11, Rab14, and Rab34 correlated with various sperm motility patterns, motion kinematics after capacitation. Moreover, Rab4 and Rab34 were associated with capacitation status before capacitation, and Rab3A, 25, and 27A correlated with capacitation status after capacitation. This is the first study to analyze the correlation between Rab proteins and sperm functions. Collectively, our results indicate that specific sperm motility and kinematics, as well as the structural condition of the sperm head and capacitation status, regulate individual Rab protein. Therefore, we expect that the current findings will be used to identify the etiology of idiopathic male infertility patients and to diagnose male fertility and that Rab proteins will be employed as biomarkers to predict and analyze male fertility.

Production of MSTN-mutated cattle without exogenous gene integration using CRISPR-Cas9.

Many genome-edited animals have been produced using clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 technology to edit specific genes. However, there are few guidelines for the application of this technique to cattle. The goal of this study was to produce trait-improved cattle using the genome-editing technology CRISPR-Cas9. Myostatin (MSTN) was selected as a target locus, and synthetic mRNA of sgRNA and Cas9 were microinjected into fertilized bovine embryos in vitro. As a result, 17 healthy calves were born, and three of them showed MSTN mutation rates of 10.5%, 45.4%, and 99.9%, respectively. Importantly, the offspring with the 99.9% MSTN mutation rate had a biallelic mutation (-12 bps) and a double-muscling phenotype. In conclusion, we demonstrate that the genome-editing technology CRISPR-Cas9 can produce genetically modified calves with improved traits.

Germline transmission of MSTN knockout cattle via CRISPR-Cas9.

Although the production of several founder animals (F0) for gene editing in livestock has been reported in cattle, very few studies have assessed germline transmission to the next generation due to the long sexual maturation and gestation periods. The present study aimed to assess the germline transmission of MSTN mutations (-12bps deletion) in MSTN mutant F0 male and female cattle. For this purpose, oocytes and semen were collected after the sexual maturation of MSTN cattle, and embryos produced by in vitro fertilization were analyzed. In addition, the embryos were subjected to additional gene (PRNP) editing using electroporation. Embryos produced by in vitro fertilization with MSTN male and female cattle were transferred to a surrogate, and one calf was successfully born. MSTN heterozygous mutation was shown by sequencing of the F1 calf, which had no health issues. As a further experiment, using electroporation, additional gene-edited embryos fertilized with the MSTN male sperm showed a high mutation rate of PRNP (86.2 ± 3.4%). These data demonstrate that the cattle produced through gene editing matured without health issues and had transmitted MSTN mutation from the germ cells. Also, additional mutation of embryos fertilized with the MSTN male sperm could enable further mutagenesis using electroporation.

Investigation of high correlation with carcass traits of SNPs of the PLCB1, C/EBPα, and TDRKH genes and the combinations of SNPs using the MDR method in the Hanwoo.

BACKGROUND: Recently, many researchers focus on the best way to produce high-quality meat, as the trend in food consumption today is to focus on quality. In general, consumers' preferences in beef differ depending on taste and meatiness. Therefore, researchers are interested in how the marbling score affects the flavors of meat or the various factors that make up the meatiness to captivate the consumers' tastes. OBJECTIVE: This study identifies single nucleotide polymorphisms (SNPs) or gene combinations that affect the carcass traits of Korean cattle (Hanwoo) by using the multifactor dimensionality reduction (MDR) method. METHODS: We collected the candidate SNPs to identify SNPs related to marbling scores from whole-exome sequencing and bovine SNP genotyping data. Using 96 Hanwoo samples, we performed PCR amplification to investigate the polymorphism status. In addition, we investigated genetic relationships between carcass traits and SNPs using 612 Hanwoo samples. Furthermore, each candidate SNP genotype and the combinations of SNP genotypes were verified to improve the accuracy of genetic relationships using MDR method. RESULTS: Twenty-four candidate SNPs associated with carcass trait and marbling scores were identified from SNP genotyping and whole-exome sequencing. Among them, three SNP markers (c.459 T > C of the PLCB1 gene, c.271 A > C of the C/EBPα gene, and g.17257 A > G of the TDRKH gene) were showed statistically significant differences between intramuscular fat and genotypes. Especially, two candidate SNPs, including c.459 T > C located in the PLCB1 gene and c.271 A > C located in the C/EBPα gene, could be highly associated with the intramuscular fat of Hanwoo quality grade. In addition, the combination of SNP genotypes is showed higher significant differences with carcass weight, backfat thickness, and longissimus dorsi muscle area. CONCLUSION: Three SNP genotypes and the combination of SNP genotypes in the PLCB1, C/EBPα, and TDRKH genes may be useful genetic markers for improving beef quality.

Change in plasma sirtuin 1 level by injection into uterus of resveratrol in Korean cattle.

Bovine embryonic development is closely associated with mitochondrial biogenesis, which is regulated by the sirtuin 1 (SIRT1). Resveratrol, which is a type of natural phenol produced by several plants and used as a dietary supplement, is the activator of SIRT1. Although it has been reported that resveratrol increased SIRT1 level in in vitro bovine blastocysts, there are no in vivo reports on the change in the plasma SIRT1 level in cows. Therefore, we investigated the change in the level of plasma SIRT1 by injecting different concentrations of resveratrol into the uterus of Korean cattle heifer. The level of plasma SIRT1 in the 1.0 µM resveratrol-injected group was the highest among all groups (P < 0.05). Although the level of plasma SIRT1 increased on days 7, 9, and 14 in the resveratrol-injected group, the level of plasma SIRT1 in the control group decreased. When 1.0 µM resveratrol was directly injected into the uterus of cows during artificial insemination, a pregnancy rate was 21.0% higher than that in the control group. In conclusion, our results identified that the level of plasma SIRT1 was increased by direct injection of resveratrol and improved conception rate by injection into uterus of cow during artificial insemination.

Detrimental effects of temephos on male fertility: An in vitro study on a mouse model.

The World Health Organization recommends temephos as a nonsystemic organophosphorus pesticide due to its low mammalian toxicity compared with other chemical compounds. Although several studies have reported that temephos may be toxic under certain conditions, little research effort has been made to evaluate its effects on mammalian fertility. Therefore, the present study was designed to evaluate the effect of temephos on sperm functions and male fertility. Initially, cauda epididymis from mouse spermatozoa was incubated with temephos (0, 0.1, 1, 10, and 100 µM). Then, sperm motility and motion kinematics, capacitation status, intracellular adenosine triphosphate level, lactate dehydrogenase level, protein kinase A (PKA) activity, and degree of tyrosine phosphorylation were analyzed. Finally, the rates of fertilization and early embryonic development were evaluated. Sperm motility and motion kinematics were found to be significantly altered in temephos groups. In addition, the acrosome reaction and capacitation significantly increased and decreased in the 100 µM temephos group, respectively. Intracellular adenosine triphosphate levels significantly decreased in the 1, 10, and 100 µM temephos groups compared with that in the control group. Moreover, PKA activity and tyrosine phosphorylation significantly decreased in most temephos groups. Further, the rates of fertilization and early embryonic development significantly decreased in all temephos groups. Taken together, it was determined that temephos had harmful effects on male fertility. Therefore, the reproductive toxicity of temephos should be considered before its use.