Revolutionary advancements, such as the reduction in DNA sequencing costs and genome editing, have transformed biotechnology, fostering progress in manipulating biomolecules, engineering cells, and computational biology. Agriculture and food production have significantly benefited from tools like high-throughput microarrays, accelerating the selection of desired traits. Genetic engineering, especially utilizing genome editing, facilitates precise alterations in plants and animals, harnessing microbiomes and fostering lab-grown meat production to alleviate environmental pressures. The emergence of new biotechnologies, notably genome editing, underscores the necessity for regulatory frameworks governing LM (living modified) organisms. Global regulations overseeing genetically engineered or genome-edited (GE) organisms, encompassing animals, exhibit considerable diversity. Nonetheless, prevailing international regulatory trends typically exclude genomeedited plants and animals, employing novel biotechnological techniques, from GMO/ LMO classification if they lack foreign genes and originate through natural mutations or traditional breeding programs. This comprehensive review scrutinizes ongoing risk and safety assessment cases, such as genome-edited beef cattle and fish in the USA and Japan. Furthermore, it investigates the limitations of existing regulations related to genome editing in Korea and evaluates newly proposed legislation, offering insights into the future trajectory of regulatory frameworks.

Background: Single nucleotide polymorphisms (SNPs) are widely used genetic markers with applications in human disease diagnostics, animal breeding, and evolutionary studies, but existing genotyping methods can be labor-intensive and costly. The aim of this study is to develop a simple and rapid method for identification of a single nucleotide change. Methods: A modified Polymerase Chain Reaction Amplification of Multiple Specific Alleles (PAMSA) and high resolution melt (HRM) analysis was performed to discriminate a bovine polymorphism in the NCAPG gene (rs109570900, 1326T > G). Results: The inclusion of tails in the primers enabled allele discrimination based on PCR product lengths, detected through agarose gel electrophoresis, successfully determining various genotypes, albeit with some time and labor intensity due to the use of relatively costly high-resolution agarose gels. Additionally, high-resolution melt (HRM) analysis with tailed primers effectively distinguished the GG genotype from the TT genotype in bovine muscle cell lines, offering a reliable way to distinguish SNP polymorphisms without the need for time-consuming AS-PCR. Conclusions: Our experiments demonstrated the importance of incorporating unique mismatched bases in the allele-specific primers to prevent cross-amplification by fragmented primers. This efficient and cost-effective method, as presented here, enables genotyping laboratories to analyze SNPs using standard real-time PCR.

Traf4 (Tumor necrosis factor Receptor Associated Factor 4) is a member of the tumor necrosis factor receptor (TNFR) - associated factors (TRAFs) family. TRAF4 is overexpressed in tumor cells such as breast cancer and associated with cytoskeleton and membrane fraction. Interestingly, TRAF4 was localized with tight junctions (TJs) proteins including OCLN and TJP1 in mammary epithelial cells. However, the expression patterns and biological function of Traf4 were not examined in preimplantation mouse embryos although Traf4-deficient mouse showed embryonic lethality or various dramatic malformation. In this study, we examined the temporal and spatial expression patterns of mouse Traf4 during preimplantation development by qRT-PCR and immunostaining, and its biological function by using siRNA injection. We found upregulation of Traf4 from the 8-cell stage onwards and apical region of cell – cell contact sites at morula and blastocyst embryos. Moreover, Traf4 knockdown led to defective TJs without alteration of genes associated with TJ assembly but elevated p21 expression at the KD morula. Taken together, Traf4 is required for TJs assembly and cell proliferation during morula to blastocyst transition.

Recent studies showed that tight junctions (TJs) integrity and assembly are required for blastocyst development in mouse and pig models. However, the biological functions of TJs associated with embryo implantation and maintenance of pregnancy were not investigated yet. To examine whether disrupted TJs affect further embryo development, we employed RNAi approach and inhibitor treatment. The embryos were injected with Cxadr (Coxsackievirus and adenovirus receptor) siRNA for knock down (KD) and treated with Adam10 (A Disintegrin and Metalloproteinase specific inhibitor 10; GI254023X; SI). We compared blastocyst development and paracellular sealing assay using FITC dextran uptake between control and KD or SI embryos. Finally, we transferred control and Cxadr KD or Adam 10 SI treated blastocyst to uteri of recipients. Cxadr KD and Adam 10 SI showed lower blastocyst development and more permeable to FITC-dextran. Moreover, we observed that half of KD and inhibited embryos failed to maintain pregnancies after the second trimester. Our findings suggested that TJs integrity is required for the maintenance of pregnancy and can be used as a selective marker for the successful application of assisted reproduction technologies.

Establishment of the Adherens junction (AJ) and Tight junction (TJ) are important steps in terms of morphological formation during preimplantation develoment. Particularly, TJ complex is crucial for cavitation in blastocyst. So far, many TJ protein/genes are revealed. However, the biological function and regulation of TJ were not elucidated during post implantation. We depleted several TJ and TJ associated genes using RNA interference, and examined preimplantation development with TJ. We tested functionality of paracellular sealing to determine integrity of TJ formation and examined TE differentiation indirectly using outgrowth assay in vitro. We observed defect of paracellular permeability in the TJ related genes knockdown(KD) blastocyst and abnormal outgrowth. Particularly, trophoblast cells were not stretched out in the KD groups. Finally, we did embryo transfer using the TJ genes KD and control blastocysts into surrogate mothers. We found lower of the implantation rates/ maintenance of pregnancy in the TJ KD groups (less than 40%) than in the controls (about 80%). In conclusion, TJ integrity is can be used as a selective marker for developmentally competent embryos and successful pregnancy.

The Korean native cattle, Hanwoo, is the most popular breed of beef cattle in Korea. However, the reproductive performance data are limited although reproduction is one of the most economically and biologically important in beef production. Therefore, this study was undertaken to investigate reproductive performance parameters including calving interval, parity for life time production. Data collected from 206,827 calvings were analyzed. There were no significant differences in calving interval and gestation days as parity increased from 2nd and 13rd parity cow, from spring to winter. However, we found a dramatic increase in calving interval after year 2000. About 1 month were increased per year ( y = 30.578x + 344.45 R² = 0.9157). Interestingly, we observed that parities for life time can be affected by birth weight. Calves with 23 kg at birth showed highest parities, 3.4±2.0 times. In summary, this study provides valuable data on reproductive performance of Hanwoo and the data presented here can be used as a standard target for optimising and enhancing reproductive performance.

Wholesale beef price is the critical factor for determining Korean native cattle, Hanwoo, farm’s income in short-term. Wholesale beef price has seasonality due to high demand in Korean traditional holidays such as Korean thanksgiving day and lunar new year’s day. Therefore, it is important to make reproduction and marketing plans for Korean Hanwoo farmers, in order to increase their farm income. However, there is no study available on changes in the expected farm income depending on reproduction and marketing schedules. This study analyzed the expected farm income per head depending on the monthly-based marketing schedules. The analysis was conducted based on the seasonality of wholesale beef price, reproduction efficiency, operating costs, relationship between carcass grade and slaughter age. The result shows that slaughter Hanwoo at the age of 29 months-old in August and January generating the highest expected farm income per head.

The aim of the present study was to evaluate and estimate timing of artificial insemination (AI) in Hanwoo heifer (Korean native cattle) that is the most popular breed of beef cattle in Korea. To determine changes in body weight of heifers around AI, body weight were measured at different stages either before or after AI. We found that daily body weight gain was higher in the pregnant cows after AI. We also investigate correlation between body mass measured by shoulder height and body length, and conception rates, used (body length+ height)2 instead of height2 for body mass index (BMI), and found that relatively more BMI heifers (>55) showed higher conception rates. Finally, we estimated body weight by measuring should height (SH), heart girth (HG), and body length (BL); BW=3.93372*HG-2.90985*SH-0.021*BL. In addition, we observed that HG is most closely correlated with BW; y(BW) = 1.77355*x(HG), R² = 0.98881. In summary, we can determine the best timing of AI using body measurement and its application including BMI.

Transcription factor called activating enhancer binding protein 2C (AP2-gamma) is found in a variety of species and expressed from oocyte stage onwards, particularly restricted to the trophectoderm. Recent studies demonstrated that ablation of Tfap2c led to failure of tight junction biogenesis, particularly the knock-down embryos of Tfap2c did not form cavity from morula to blastocyst in mouse and pig. We speculated that the Tfa2pc may also be involved in desmosome biogenesis because blastocoel formation is coincident with the establishment of desmosome. To determine this, we depleted Tfap2c injecting siRNA into one-cell zygote and analysed the expression levels of genes that are required for desmosome complex such as PkP2, Pkp3, Dsc2, and Dsg2. We found only Pkp3 was up-regulated in the knockdowned morula embryos. Interestingly, upstream region of Pkp3 had putative Tfap2c binding sites. In conclusion, our results suggest that Tfap2c is not a crucial factor but somehow it might be involved in desmosome biogenesis directly or indirectly via Pkp3.

Calculating break-even price of calf production is closely associated with reproductive efficiency. To determine the price, we need data from reproduction records including number of claves weaned, number of cows exposed for breeding, and annual cash coast per cow, and average weaning or market weight of claves sold and retained. Unfortunately, the data were not available in Korea native cow (Hanwoo). To evaluate the performance and the price, we collected calving interval from about 60,000 cows for last 10 years and estimated reproductive performance. Calving interval was increased 4.3% and pregnancy rate was decreased about 1.4∼2.8% year-on-year. Increases in growth rates of number of cow and semen per calf supported the low reproductive performance. Finally, break-even price was calculated using estimated percent calf crop and demonstrated that growth rate of break-even price is larger than that of annual cash per cow, suggesting cow-calf profitability and financial efficiency in Korea native cow (Hanwoo) is getting worse.

Successful somatic cell nuclear transfer (SCNT) has been reported across a range of species using a range of recipient cells including enucleated metaphase II (MII) arrested oocytes, enucleated activated MII oocytes, and mitotic zygotes. However, the frequency of development to term varies significantly, not only between different cytoplast recipients but also within what is thought to be a homogenous population of cytoplasts. One of the major differences between cytoplasts is the activities of the cell cycle regulated protein kinases, maturation promoting factor (MPF) and mitogen activated protein kinase (MAPK). Dependent upon their activity, exposure of the donor nucleus to these kinases can have both positive and negative effects on subsequent development. Co-ordination of cell cycle stage of the donor nucleus with the activities of MPF and MAPK in the cytoplast is essential to avoid DNA damage and maintain correct ploidy. However, recent information suggests that these kinases may also effect reprogramming of the somatic nucleus and preimplantation embryo development by other mechanisms. This article will summarise the differences between cytoplast recipients, their effects on development and discuss the potential role/s of MPF and or MAPK in nuclear reprogramming.

The vast majority of embryo generated by Assisted Reproductive Technologies (ART) do not result in a live offspring and a multiple birth is the single biggest health risk associated with human fertility treatment, and the used of frozen embryos increased for medical or personal reasons. However, practical and ethical reasons might hamper study of human embryos. Therefore, animal models are necessary to elucidate the molecular and morphological changes during development. In the serial experiments, we employed mouse embryos and a Cdx-inducible ES cell system that transdifferentiates into TS cells. We found aberrant gene expression profiles including apoptosis associated (Bcl2), lineage formation related genes (Cdx-2, Tcfap2c, Oct4, and Nanog), and/or mitochondrial DNA replication related genes (mt-cox-1, mt-cox-2, Polg, Polg2, Tfam) in mouse embryos that showed developmentally retardation between morula to blastocyst transition or post implantation development after embryo transfer to surrogate mothers, compared to control embryos. To determine direct interaction between knockdown genes via siRNA approach and putative down-stream genes involved in blastocyst formation and further development, we carried out qPCR and Chip assay in either mouse embryos or the ES cells. qPCR and Chip assay results showed target gene directly bound to promoter regions of down-regulated genes in TS cells. In conclusion, we suggested that an increased understanding of epigenetic regulation of early embryonic development through animal models may ultimately lead to better methodologies for selecting more competent embryos and and/or protocols for augmenting embryos viability.