黄曲霉毒素分解酶基因克隆及其在大肠杆菌中的融合表达

通过GenBank公布的黄曲霉毒素分解酶基因序列,合成ADTZ基因引物。从发霉的玉米样品中提取混合菌液,经菌液PCR扩增,筛选ADTZ基因片段,构建表达载体,将目的基因转化到大肠杆菌中,并进行诱导表达。通过SDS-PAGE检测表达产物大小和浓度。通过酶解试验检测表达产物对AFB1的降解能力。结果成功从发霉玉米混菌悬液中扩增到ADTZ基因,该基因序列全长2 088 bp,与已报道的ADTZ基因相似度达到99%。该基因可在大肠杆菌中进行融合表达,表达产物蛋白大小为118.5 kDa。重组大肠杆菌培养后获得的粗酶液可降解发霉玉米中的AFB1,降解率达到77.69%。

Cloning of aflatoxin-detofizyme gene and fusion expression in Escherichia coli

The ADTZ gene primers were synthesized through the ADTZ gene sequence published by GenBank. The ADTZ gene fragment was screened by PCR from the mixed bacterial solution which extracted of the moldy corn sample, and transformed into E. coli by constructing expression vector. The size and concentration of the ADTZ expression product were detected by SDS-PAGE. The degradation ability of the expression product to aflatoxin AFB₁ was tested by enzymatic hydrolysis. The results showed that the ADTZ gene was successfully amplified from the moldy maize suspension. The full length of the ADTZ gene sequence was 2 088 bp, and the similarity with the reported ADTZ gene was 99%. The gene can be fused and expressed in E. coli. The protein size of the expressed product was 118.5 kDa. The crude enzyme solution of recombinant E. coli could degrade AFB₁ in mildewed maize, and the degradation rate reached 77.69%.