Induced Expression and Optimal Expression Conditions of Recombinant Alpha-bungarotoxin Gene Fusion Protein

[Objective] This study aimed to obtain recombinant alpha-bungarotoxin （a-BG-0 gene fusion protein with biological activity and investiagte its fusion expression. [Method] The plasmid pGEX-a-BGT was transformed into E coil BL21 （DE3） and BL21 （DE3） plysS host bacteria to identify the optimal engineering strain. Fusion expression of the optimal engineering strain was induced, in order to optimize the induced expression conditions of the soluble fusion protein. [Result] JP-a-BGT was identified as the optimal engineering strain, which could express fusion protein after induced by IPTG. The optimal induced expression conditions of the soluble fusion protein were investigatect JP-a-BGT was incubated at 37 ℃ for 2.5 h and induced with 0.50 mmol4. IPTG for 4 h at 22 ℃, and the expression level of the soluble fusion protein reached 18.42%. [Conclusion] This study laid a solid foundation for the subsequent purification of fusion proteins and the separation and purification of a-BGT.

Optimization of Expression Conditions for Immunodominant Membrane Protein Gene of Phytoplasmas in E.coli

[Objective] This study aimed to increase the expression level of immun- odominant membrane protein gene （Imp） of phytoplasmas in E. coil BL21 （DE3）. [Method] On the basis of orthogonal experiment, effects of different culture conditions on recombinant bacteria E. coil BL21-pET-28a（＋）-Imp were investigated. Based on the obtained optimal culture condition, effects of different induction conditions on the ex- pression level of Imp protein were explored. The expression level of Imp fusion pro- tein was analyzed by using SDS-PAGE and Gene Tools software. [Result] The re- sults showed that the optimal conditions for culture were at 37℃, pH 7.0, with liq- uid volume of 20% and oscillation speed of 200 r/min, for induction were at 37℃ for 6 h, with initial OD600 of about 1.5 and IPTG final concentration of 0.1 mmol/L. [Conclusion] The expression level of Imp achieved 70.98 mg/L under the optimal conditions. Optimized conditions for expression of Imp fusion protein in E. coil were determined.

Expression of ChIFN-α in Pichia pastoris and Optimization of Its Expression Condition

Because yeast codon preferred to synthesize the gene sequence of ChIFN-α mature peptide, Pichia pastoris expression system was connected into vector pPICgK, to construct recombinant plasmid pPIC9K-α. After digestion and linearization, the recombinant vector was cloned into P. pastor/s GS115, high copy transformant and its methanol utilization type were screened, and its expression conditions were optimized. The results showed that ChlFN-α successfully expressed in P. pastor/s, the expression amount of recombinant protein was the largest after induced expression in 0.5 % methanol containing 2% acid hydrolyzed casein at 28 ℃ for 72 h, and its antiviral activity was the highest.

Conditional gene expression in invertebrate animal models

A mechanistic understanding of biology requires appreciating spatiotemporal aspects of gene expression and its functional implications.Conditional expression allows for (ir)reversible switching of genes on or off,with the potential of spatial and/or temporal control.This provides a valuable complement to the more often used constitutive gene (in)activation through mutagenesis,providing tools to answer a wider array of research questions across biological disciplines.Spatial and/or temporal control are granted primarily by(combinations of) specific promoters,temperature regimens,compound addition,or illumination.The use of such genetic tool kits is particularly widespread in invertebrate animal models because they can be applied to study biological processes in short time frames and on large scales,using organisms amenable to easy genetic manipulation.Recent years witnessed an exciting expansion and optimization of such tools,of which we provide a comprehensive overview and discussion regarding their use in invertebrates.The mechanism,applicability,benefits,and drawbacks of each of the systems,as well as further developments to be expected in the foreseeable future,are highlighted.