[Construction of a replicative expression vector based on the porcine circovirus 2 replicon].

The antigen gene expression level of a DNA vaccine is the key factor influencing the efficacy of the DNA vaccine. Accordingly, one of the ways to improve the antigen gene expression level of a DNA vaccine is to utilize a plasmid vector that is replicable in eukaryotic cells. A replicative DNA vaccine vector pCMVori was constructed based on the non-replicative pcDNA3.1 and the replicon of porcine circovirus 2 (PCV2) in this study. An EGFP gene was cloned into pCMVori and the control plasmid pcDNA3.1. The two recombinant vectors were transfected into PK-15 cell, and the plasmid DNA and RNA were extracted from the transfected cells. Real-time PCR was used to determine the plasmid replication efficiency of the two plasmids using plasmid before and after Bcl Ⅰ digestion as templates, and the transcription level of the Rep gene in PCV2 replicon was detected by RT-PCR. The average fluorescence intensity of cells transfected with the two plasmids was analyzed with software Image J, and the transcription level of EGFP was determined by means of real-time RT-PCR. The results showed that the replication efficiency of pCMVori in PK-15 cells incubated for 48 h was 136%, and the transcriptions of Rep and Rep' were verified by RT-PCR. The average fluorescence intensity of the cells transfected with pCMVori-EGFP was 39.14% higher than that of pcDNA3.1-EGFP, and the transcription level of EGFP in the former was also 40% higher than that in the latter. In conclusion, the DNA vaccine vector pCMVori constructed in this study can independently replicate in eukaryotic cells. As a result, the expression level of cloned target gene was elevated, providing a basis for developing the pCMVori-based DNA vaccine.

Effects of Schisandra chinensis Polysaccharide-Conjugated Selenium Nanoparticles on Intestinal Injury in Mice.

Schisandra chinensis polysaccharide (SCP) is an experimental therapeutic for the treatment of intestinal injury. Selenium nanoparticle modification can improve the bioactivity of polysaccharides. In this study, SCP was firstly extracted and purified by a DEAE-52 column, then SCP-Selenium nanoparticles (SCP-Se NPs) were prepared, and the procedure was optimized. Thereafter, the obtained SCP-Se NPs were characterized by transmission electron microscope, X-ray diffraction, energy-dispersive X-ray spectroscopy, and Fourier transform infrared spectroscopy. The influence of different storage environments on the stability of colloidal SCP-Se NPs was also investigated. Finally, the therapeutic effects of SCP-Se NPs on LPS-induced intestinal inflammatory injuries in mice were evaluated. Results showed that the optimized SCP-Se NPs were amorphous, uniform, spherical particles with a diameter of 121 nm, and the colloidal solution was stable at 4 °C for at least 14 d. Moreover, SCP-Se NPs could more effectively alleviate LPS-induced diarrhea, intestinal tissue injury, and tight junction destruction and decrease the elevated expression levels of TNF-α, IL-1ß, and IL-6 compared with SCP. These results demonstrate that SCP-Se NPs may alleviate LPS-induced enteritis through their anti-inflammatory effects, indicating that SCP-Se NPs can serve as a good candidate for preventing and treating enteritis in the livestock and poultry industry.