Objective:To construct a recombinant adenovirus expressing the non-structure protein 4(NSP4) of A group rotavirus, and evaluate its immune response effects. Methods: NSP4 gene from wildtype rotavirus was cloned into a shuttle plasmid (pshuttle-CMV). The pshuttle-NSP4 is transformed into E.coli BJ5183 to make a homologous recombination with adenovirus skeleton genome(pAdeasy). And the recombinant plasmid pAd-NSP4 is transfected into Ad-293 cell to produce mature adenovirus particles. ICR mice are vaccinated with this recombinant adenovirus by intramuscular and intranasal administration. Results: The recombinant adenovirus titer rise to 108.25CCID50/ml after 3 time passaged. The transcription and expression of NSP4 can be determined by RT-PCR and immunofluorescene assay in vitro. In intramuscular and intranasal administration group, ELISA determines specific serum antibody mean titer at 1:320 and 1:1436.8 level; neutralizing antibody titer at 1:45.3 and 1:71.8. Conclusion: The recombinant adenovirus containing rotavirus NSP4 gene can induce effective immune response against rotavirus. Intranasal administration produce a better protective efficacy than intramuscular in mice. It may be a good candidate to develop a novel engineering vaccine in the future.
Resveratrol is an important plant stilbene with considerable pharmaceutical values. To achieve the de novo biosynthesis of resveratrol in E. coli, a heterologous resveratrol biosynthetic pathway consisting of tyrosine ammonia lyase (TAL), coumaroyl-CoA synthase (4CL) and stilbene synthase (STS) was constructed. The engineered strain only produced 2.67 mg/L resveratrol after 3 days cultivation. To improve the efficiency of heterologous pathway, the 4CL and STS modules were further engineered using strategies of fusion expression, high-copy expression and promoter engineering. Eventually, the yield of resveratrol reached 25.76 mg/L with a 9.6-fold improvement compared with the initial strain. The useful information for the construction of more efficient recombinant resveratrol producer was provided, and it laid a foundation for the large-scale production of resveratrol through microbial fermentation.
Circulating microRNAs (miRNAs) are a family of important small non-coding RNA molecules, which participate in communication between different cells. Because circulating miRNA can be used as biomarkers of many diseases, it has attracted more attention recently. The sources and existence forms of circulating miRNAs were briefly summarized, sample preparation and detection methods of circulating miRNAs were analyzed. Additionally, the applications of circulating miRNAs were also discussed. It can provide references for further study and use of miRNAs in theory and practice fields.
