苏云金杆菌ZLL13晶体蛋白分析及其菌糠液态培养基的优化

Analysis of Crystal Protein of Bacillus thuringiensis ZLL13 and Optimization of Liquid Culture Medium of Spent Mushroom Substrate

食用菌栽培废料,简称菌糠(spent mushroom substrate, SMS)是食用菌栽培和生产的残留物,其含有丰富的甲壳素、木质纤维和蛋白质等,可为苏云金芽孢杆菌(Bacillus thuringiensis, Bt)的生长提供所需的营养物质。本研究以优化后的前处理条件制备的菌糠浸提液(2%硫酸, 121℃, 1 h)作为主要碳源,通过单因素试验、Plackett-Burman设计、最陡爬坡和响应面分析等方法来优化最佳培养基组分,结果表明,54%SMS浸提液,31.9 g/L大豆饼粉、0.88 g/L CaCO3、0.4 g/L MnSO4、0.5 g/L K2HPO4和0.4 g/L吐温100为最佳培养基配方,且优化后培养基(1.8×108/mL)产生的孢子数是原始SMS培养基(0.065×108/mL)的27倍,这不仅为菌糠的二次利用提供一种新的有效方法,而且也可以大大降低生产Bt所需要的发酵成本,具有良好的应用前景。

Spent mushroom substrate(SMS) has been demonstrated that the entomotoxicity and chitinase activity of Bacillus thuringiensis based biopesticides may be increased by fortified with chitin during production. To develop a cost-effective biopesticide, spent mushroom substrate extract was studied as a potential chitin and carbon source for cultivating Bacillus thuringiensis. Subsequently, Spent mushroom substrate extract(2% H2 SO4, 121℃, 1 h)was selected for BtiZLL13 fermentation, Plackett-Burman design(PB) and response surface methodology(RSM)were employed to optimize the medium components for production of Bti-based biopesticide. The optimal composition was as followes: 54% SMS extract, 31.9 g/L soybean meal, 0.88 g/L CaCO3, 0.4 g/L MnSO4, 0.5 g/L K2 HPO4, and 0.4 g/L Tween 100. Under the optimized conditions, more than 27 times improvement in production of spores was achieved compared with the original SMS medium. Results from this study suggest that SMS is a highly cost-effective raw material for mass production of Bti.