Construction of Expression Vector for Porcine Gastrin-releasing Peptide Fusion Protein

[Objectives]This study was conducted to obtain porcine gastrin-releasing peptide(GRP)fusion protein.[Methods]The constructed pET32a(+)-GRP plasmid was transformed into Escherichia coli BL21(DE3)competent cells to obtain the pET32a(+)-GRP-BL21(DE3)fusion protein expression strain,which was induced with 0.5 mM IPTG at 25℃and 150 r/min for 12 h,and the His-tagged GRP fusion protein was detected by SDS-Page gel electrophoresis and Western Blot.[Results]After optimizing the IPTG-induced expression conditions,it was confirmed that the porcine GRP fusion protein was obtained,and the porcine GRP fusion protein was soluble,stable and highly active.[Conclusions]This study lays a foundation for the subsequent preparation of anti-pig GRP antibodies.

Effects of LPS on the Gene Expression of NMB and Its Receptor in the Hypothalamic-pituitary-testicular Axis of Rabbits

[Objectives]This study was conducted to investigate the effect of lipopolysaccharide(LPS)on the gene expression levels of neuromedin B(NMB)and its receptor(NMBR)in the hypothalamic-pituitary-testis(HPT)axis of male rabbits.[Methods]Forty male New Zealand white rabbits were randomly divided into an experimental group injected with LPS and a control group.At 0,1.5,3,6 and 12 h after LPS injection,four rabbits in each group were sacrificed,and the hypothalamus,pituitary and testicular tissues were collected.RT-PCR was carried out to detect the expression of rabbit NMB and NMBR mRNA in the HPT axis.[Results]NMB and NMBR mRNA were expressed in the hypothalamus,pituitary gland and testis of rabbits,and there was a certain time effect,indicating that the NMB/NMBR system is involved in the activities of the hypothalamus-pituitary-testis(HPT)axis.At 1.5 h after LPS stimulation,the expression of NMB mRNA in the hypothalamus increased significantly,and showed an extremely significant difference from the control group(P<0.01),while the expression of NMBR mRNA in the hypothalamus increased,but was not significantly different from the control group(P>0.05).The expression levels of NMB and NMBR mRNA in the pituitary and testis tissues in the HPT axis at different time after the LPS treatment were not significantly different from those in the control group(P>0.05).Therefore,LPS stimulation still has a certain effect on the expression of NMB and its receptor mRNA in the HPT axis.[Conclusions]This study is of great significance for the in-depth study on the mechanism of damage caused by immune stress to livestock and poultry reproduction.