The aim of the present study was to investigate the beneficial effect of polydatin (PD), the glycoside form of resveratrol, on embryo development in vitro. Oocytes were aspirated from ovaries of Korean Hanwoo cows and cultured until Day 8 in a humidified atmosphere of 5% CO in air at 38.5°C. Protein and gene expression levels were determined through confocal microscopy and reverse transcription-polymerase chain reaction respectively, whereas the number of total and apoptotic cells in Day 8 blastocysts was determined using Hoechst 33342 staining and terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling. Of the different concentrations of PD (0.5, 1.0 and 2.0µM) added to the IVM medium, only 1.0µM PD significantly improved blastocyst development. Immunofluorescence analysis confirmed that protein levels of sirtuin 1 (Sirt1) increased significantly (P<0.05) after PD treatment, whereas levels of reactive oxygen species (ROS) were significantly (P<0.05) decreased, as evidenced by reductions in 8-oxoguanine immunoreactivity. Similarly, protein levels of nuclear factor (NF)-κB and cyclo-oxygenase (COX)-2 were significantly (P<0.05) lower in the PD-treated group than in the control group. Treatment with 1.0µM PD reduced gene expression of BCL2-associated X protein, inducible nitric oxide synthase, COX2 and Nfkb, but increased the expression of Sirt1, supporting the immunofluorescence data. PD possesses antioxidant activity and is useful for embryo development in vitro. We conclude that supplementation of IVM medium with PD improves embryo developmental competence via Sirt1.

Pleurotus spp. have been used for edible and medicinal purposes in Asian countries for a long time. The fruiting bodies of the Pleurotus ostreatus, Pleurotus citrinopileatus and Pleurotus salmoneostramineus contained many physiologically beneficial substances for human health. Therefore, it is necessary to study the genetic diversity of Pleurotus mushroom cultivars commercially cultivated in Korea. Eleven strains of Pleurotus spp. were collected from different geographical regions in South-East Asia and ITS regions of rDNA and RAPD of genomic DNA were analyzed. The size of the ITS1 and ITS2 regions of rDNA from the different strains varied from 167 to 254 bp and 156 to 213 bp, respectively. The sequence of ITS1 was more variable than that of ITS2, and the 5.8S sequences were identical. A phylogenetic tree based on the ITS region sequences indicated that selected strains could be classified into 4 clusters. Eleven Pleurotus species were also analyzed by RAPD with 20 arbitrary primers. Ten of these primers were efficiently amplified the genomic DNA. The number of amplified bands varied with the primers and strains, with polymorphic fragments in the range from 0.1 to 2.0kb. The results revealed that genetic diversity of selected strains of P. ostreatus, P. citrinopileatus and P. salmoneostramineus is low.

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Background: Allergy health problems worldwide are mainly caused by allergens from domestic cats ( Felis catus). Fel d 1 is a major allergen that causes severe allergic reactions in humans, including rhinitis, conjunctivitis, and life-threatening asthma. The treatment for allergic reactions to Fel d 1 in humans is still in its infancy. Patients with cat allergies anticipate hypoallergenic cats. Methods: We used the CRISPR-Cas9 system, microinjection, and embryo transfer to knock out the chain 2 (CH2) genome of Fel d 1 in cats. T7 endonuclease 1 (T7E1) assay and Sanger sequencing analysis were used to confirm CH2 knockout cats. Fel d 1 level in CH2 knockout cats was assessed by enzyme-linked immunosorbent assay (ELISA). Cytoplasm injection clone technology (CICT) was used to clone the CH2 knockout cat. Results: We report the first successful generation and cloning of CH2 knockout cats using the CRISPR-Cas9 system and CICT. CH2 knockout cats were confirmed using T7E1 and Sanger sequencing. Cloned CH2 knockout cat was verified by microsatellite analysis. Remarkably, ELISA proved that Fel d 1 level of CH2 knockout cats was extremely low compared with that of wild type domestic cats. Conclusion: CH2 knockout cats we generated showed an extremely low level of Fel d 1 and could be hypoallergenic cats. Our study indicates that creating hypoallergenic cats using the CRISPR-Cas9 system is a significant step forward in the consequence that these cats can safely approach allergic patients.

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