Optimization of induced fermentation conditions for a high-yielding Bacillus subtilis of metallothionein
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摘要: 金属硫蛋白为一类广泛存在于生物体内的多功能诱导性蛋白,具有转运储存金属离子、清除自由基、激活（失活）锌调节蛋白等功能,近年来已成为研究与开发的热点。本文以一株基因重组枯草芽孢杆菌为实验对象,对其诱导表达金属硫蛋白发酵条件进行研究。结果表明,枯草芽孢杆菌最优生长条件为:培养温度37℃,培养基初始p H7,摇床转速220 r/min,接种量4%;最佳诱导发酵条件为:菌液OD600=3.9,IPTG终浓度1 mmol/L,摇床转速220 r/min,诱导时间48 h,诱导金属Cd2+终浓度为50μmol/L,发酵培养基无机盐为1%MgSO4、碳源为2%蔗糖、氮源为1.5%酵母浸出物+胰蛋白胨+氯化铵（2∶2∶1）,枯草芽孢杆菌MT表达量达到最高为0.135 mg/m L。本研究为开发天然、高效海洋源金属硫蛋白提供了一定理论依据。Abstract: Metallothionein ( MT) is a kind of multifunctional inducible protein widely distributed in the organism, with the functions of transportation and storage of metal ions, scavenging free radicals function, activation ( inactivation) function of zinc regulatory proteins, and has been one of the hot fields of biology and medicine in basic research and applied research in recent years.In this paper, the fermentation conditions of a strain of high yield recombinant Bacillus subtilis strain were optimized.Through the single factor experiment, the optimal conditions for the growth of recombinant Bacillus subtilis as follows, temperature 37 ℃, initial p H7, rotation speed 220 r/min, inoculation amount 4%, and the optimal fermentation conditions of induction as follows, OD600= 3.9, IPTG concentration 1 mmol/L, the rotate speed 220 r/min, the induction time 48 h, 50 μmol/L Cd2+, MgSO4 ( 1%) , sucrose ( 2%) , nitrogen source ( 1.5% yeast extract + peptone + ammonium chloride) ( 2 ∶ 2 ∶ 1) , the expression of MT reached 0.135 mg/m L.This study provides some theoretical basis for the development of the natural, efficient marine source metallothionein.
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Keywords:
- metallothionein /
- Bacillus subtilis /
- fermentation
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