棉田Cry1Ac蛋白降解菌株12T-103的分离鉴定及降解能力研究
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摘要
为分离鉴定棉田转Cry1Ac基因棉的Cry1Ac蛋白土壤降解细菌及测定其降解能力。采用蛋白质凝胶电泳(SDSPAGE)、酶联免疫法(ELISA)法和常规细菌培养与鉴定技术,对第一师12团棉田土壤中Cry1Ac蛋白降解菌进行分离、纯化、形态生理生化观察以及降解功能测定。结果表明:从棉田土壤分离一株具有降解Cry1Ac蛋白能力的土壤细菌,命名为12T-103菌株,细菌形态学、生理生化特征和16S r DNA序列鉴定为枯草芽孢杆菌(Bacillus subtilis),该菌株对Cry1Ac蛋白的降解效果较为明显,在1h时Cry1Ac蛋白含量由15. 95μg·L-1降至0. 68μg·L-1,其降解程度达极显著水平(p <0. 01),其降解率为92. 26%。结论:分离于转基因棉田的12T-103菌株为枯草芽孢杆菌(Bacillus subtilis),该菌株对Cry1Ac蛋白的降解能力较强。
For isolation and identification of Cry1 Ac protein degrading bacterial strain and its degradation ability from the soil of planting Bt cotton in the farm 12,first division of Xinjiang production and construction corps,the Cry1 Ac protein degrading bacterial strain and its bacterial form,physiology and biochemistry in the soil were isolated,purified and detected with SDS-PAGE,enzyme linked immunoassay( ELISA) and conventional bacterial culture and identification technologies. The results showed that one Cry1 Ac protein degrading bacterial strain was isolated and named as 12 T-103,and the 12 T-103 was identified as Bacillus subtilis according to the phenotype observation,characteristics of physiology and biochemistry and 16 S r DNA analysis,which had great significant effect on the degradation of Cry1 Ac protein from 15. 95μg·L-1 to 0. 68μg·L-1 in 1 h( p < 0. 01),and the degradation rate of Cry1 Ac protein was 92. 26%. It indicated that the 12 T-103 from the Bt cotton planting soil was Bacillus subtilis,and had strong degradation ability on Cry1 Ac protein.
For isolation and identification of Cry1 Ac protein degrading bacterial strain and its degradation ability from the soil of planting Bt cotton in the farm 12,first division of Xinjiang production and construction corps,the Cry1 Ac protein degrading bacterial strain and its bacterial form,physiology and biochemistry in the soil were isolated,purified and detected with SDS-PAGE,enzyme linked immunoassay( ELISA) and conventional bacterial culture and identification technologies. The results showed that one Cry1 Ac protein degrading bacterial strain was isolated and named as 12 T-103,and the 12 T-103 was identified as Bacillus subtilis according to the phenotype observation,characteristics of physiology and biochemistry and 16 S r DNA analysis,which had great significant effect on the degradation of Cry1 Ac protein from 15. 95μg·L-1 to 0. 68μg·L-1 in 1 h( p < 0. 01),and the degradation rate of Cry1 Ac protein was 92. 26%. It indicated that the 12 T-103 from the Bt cotton planting soil was Bacillus subtilis,and had strong degradation ability on Cry1 Ac protein.
引文
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[4]张美俊,段云,杨武德,等.转Bt基因棉粉碎叶还土对土壤微生物活性的影响[J].山西农业大学学报(自然科学版),2013,33(2):93-97.
[5]赵辉欣,刘阳,邢福国. Cry1Ac蛋白降解菌株FJSB3的分离鉴定及降解特性[J].核农学报,2011,25(5):922-926.
[6]赵辉欣.转基因水稻秸秆中Cry1Ac蛋白降解的研究[D].中国农业科学学院,2011.
[7]李良秋,郑贺云,蔡卓平,等.新疆阿克苏盐碱地土壤细菌的分离研究[J].生物技术进展,2013,4(3):281-287.
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[9]王晓云,王慧,赵燕,等.紫外诱变选育高活性蛋白酶枯草芽孢杆菌及其降解饲料能力评价[J].中国水产科学,2016,23(6):1351-1357.
[10]王斌,张腾霄,刘利军,等.枯草芽孢杆菌对金针菇发酵液蛋白质降解研究[J].食品工业,2017,38(2):193-196.
[11]郑贺云,黎志坤,李超.新疆阿克苏地区盐碱地细菌类群多样性及优势菌群分析[J].微生物学通报,2012,39(7):1031-1043.