Bt群体信号应答因子nprR基因的缺失对cry1Ac基因表达的影响

Effect of Quorum Sensing response regulator nprR deletion on expression of Cry protein in Bacillus thuringiensis

摘要:

摘要：【目的】研究群体信号应答蛋白编码基因nprR在苏云金芽胞杆菌（Bacillus thuringiensis，Bt）HD-73菌株晶体蛋白形成过程中的作用。【方法】通过同源重组，构建了HD-73 nprR基因缺失突变菌株HD73（ΔnprR ）。利用启动子-lacZ融合、SDS-PAGE方法，测定不同培养基中nprR基因转录活性及nprR基因缺失对cry1Ac转录及表达的影响。【结果】启动子转录活性分析表明，在LB和SSM培养基中nprR基因从对数期结束（T0）开始表达，稳定期持续表达。在LB培养基中，nprR基因的缺失使cry1Ac基因在生长过渡期和稳定期前期转录活性显著提高，同时HD73（ΔnprR ）菌株Cry蛋白生成量也明显高于出发菌株HD-73，但是在芽胞形成释放后，Cry蛋白的表达没有明显的区别。【结论】在丰富培养基中苏云金芽胞杆菌nprR基因的缺失在生长过渡期和稳定期前期能够提高cry1Ac基因转录和表达，从而缩短了cry基因表达时间，并且Cry蛋白总产量与出发菌株相当。

Abstract:

Abstract: [Objective] The role of Quroum Sensing response regulator nprR on the expression of Cry protein in B. thuringiensis HD-73 was studied. [Methods] The nprR gene deletion mutant HD73 (ΔnprR) was constructed by using of homologous recombination. Beta-galactosidase assay of cry1Ac′-lacZ gene fusion and SDS-PAGE in both HD-73 and HD73 (ΔnprR) strains were performed to analyze the effect of nprR gene deletion on expression of cry1Ac gene. [Results] Beta-galactosidase assay of nprR′-lacZ in both LB and Schaeffer’s sporulation medium showed nprR gene in B. thuringiensis was initially transcripted at T0 (end of Logarithmic growth phase) and keeping expression in stationary phase. Beta-galactosidase assay of cry1Ac′-lacZ and SDS-PAGE indicated that expression of cry1Ac gene in HD73 (ΔnprR) was stronger than that in HD-73 during transition phase and early stationary phase. However, Cry expressed product between HD-73 and HD73 (ΔnprR） in LB medium has no significant difference when crystal and spore were released. [Conclusion] The deletion of nprR increased expression and transcription activity of cry1Ac during transition and early stationary phase in rich media.