Somatic nuclear transfer technology has become increasingly promising in biomedicine and agriculture. Whereas the approach remains inefficient and underlying mechanisms remain ambiguous. Although cloned embryos have similar in vitro developmental capacity as in vitro fertilized (IVF) embryos before implantation, they appeared to have much lower full-term developmental efficiency in pig and cattle, and thus it would be reasonable to postulate that profound distinction at the molecular level should exist between them. Herein, RNA sequencing technique was used to screen differentially expressed genes in cloned and IVF blastocysts, and in total 628 differentially expressed transcripts were obtained, among which, 280 transcripts are up-regulated and 348 transcripts are down-regulated in cloned blastocysts. Moreover, one statistically significant pathway associated with endoplasmic reticulum (ER) protein processing was enriched, and some ER-stress markers such as ATF4, ATF6, PDIA3, HSPA1B, HSP40 and HSP90 between cloned and IVF blastocysts were suggested. Additionally, some developmentally important genes such as lipid metabolism related genes (MGLL, DDHD2 and FADS2) and epigenetic modification genes (DNMT1, KDM5C and MBD3L5) were found differentially expressed between cloned and IVF embryos.

Many animal feed grains contain high β-glucan in the cell wall. Pigs do not secret β-glucanase to degrade the β-glucan in their feed. The indigestible β-glucan not only blocks the release of nutrients from the grain cell wall, but also increases the digesta viscosity in the gastrointestinal tract of pigs. Therefore, dietary β-glucan significantly inhibits nutrient digestion and absorption in pigs. Transgenic expression of β-glucanase in the digestive tract of pigs may offer a solution to solve this problem. In the current study, four arti?cial codon-optimized β-glucanases genes was prepared and expressed in porcine cells. Only pBgA and pEgx showed high activity in transfected pig kidney cells. To improve the pH range and pH stability of β-glucanase, the two β-glucanases, pBgA and pEgx, were co-expressed in pig kidney cells and salivary gland cells by Linker A3 or 2A peptide. The resulting dual enzymes of pBgA3pEg and pBg2ApEg showed significantly enlarged pH range and significantly increased pH stability, as compared to parental enzymes. These results provide useful data for future study on increasing the feed digestibility of pigs by transgenic expression of β-glucanase in their salivary glands.