A Toxin Protein Quantitative Detection Method for Bacillus Thuringrensis Subsp. Israelensis

doi:10.14088/j.cnki.issn0439-8114.2019.03.026

Abstract

Abstract: To provide a quantitative detection method for toxin protein of Bacillus thuringrensis subsp. israelensis （Bti），and to improve the quality control technique. The samples of Bacillus thuringrensis subsp. israelensis were washed in water with a certain concentration of phosphoric acid buffer solution， and the residue of culture medium was removed. The bacterial toxin protein collected after centrifugation was detected by SDS-PAGE electrophoresis and the content was determined. The key problem of the determination of impurity interfering toxin protein in the sample was solved by washing the sample with phosphoric acid buffer and water. This method can accurately detect and quantify the two major molecular weight toxin proteins of Bacillus thuringrensis subsp. israelensis. The method was used to detect the subspecies of Bacillus thuringrensis subsp. israelensis. The results were reproducible， the detection error could be controlled within 5% of the average value， and there was a significant correlation between the detection results and the biological titer of the products. The method is reasonable in conception and simple in operation， and can provide reference for further product quality management.

Key words: Bacillus thuringrensis subsp. israelensis； toxin protein； SDS-PAGE gel； electrophoresis

CLC Number:

JIN Hai-yan，HU Ling-yun，LIU He-mei，ZHOU Li，HU Xiao，CHENG Zhi-guo，LI Qing，LIU Hua-mei. A Toxin Protein Quantitative Detection Method for Bacillus Thuringrensis Subsp. Israelensis[J]. HUBEI AGRICULTURAL SCIENCES, 2019, 58(3): 96-101.

References

[1] 陆宝麟. 我国疟疾及其防治问题[J].中华流行病学杂志,1982(3):279.
[2] 王逸民,葛继乾,梁雪嘉,等.北京地区三带喙库蚊乙型脑炎感染率调查[J].中华流行病学杂志,1983(4):273.
[3] 卫生部疟疾专家咨询委员会.一九九七年全国疟疾形势[J].中国寄生虫学与寄生虫病杂志,1998(16):161.
[4] 张国才,董学书,王学忠,等.腾冲县昆明按蚊的分布及传疟作用调查[J].中国寄生虫病防治杂志,1989(2):206.
[5] 徐淑惠. 中国丝虫病防治[J].中国寄生虫病防治杂志,1997(10):246.
[6] WORLD HEALTH ORGANIZATION.Geographical Distribution of Arthropod-borne Diseases and Their Principal Vectors[M].Switzerland Geneva:Division of Vector Biology and Control,1989.
[7] 陆宝麟. 中国登革热媒介及其防治[M].贵阳:贵州人民出版社,1990.
[8] IBARRA J E,FEDERICI B A J. Isolation of arelatively toxic 65-kDa protein inclution from the parasporal body of Bacillus thuringiensis subsp. iraelensis[J].Bacteriology,1986,165(2):527-533.
[9] 蒲蛰龙. 苏云金芽孢杆菌以色列变种防治蚊幼虫的研究[M].广州:中山大学出版社,1983.
[10] 吴继星. 细菌生物防治蚊子的研究进展[J].微生物学研究与应用,1993(1):27-30.
[11] 吴秋雁. 蚊幼虫感染苏云金杆菌以色列变种后的病理研究[J].昆虫学报,1986,29(1):35-40.
[12] 宋金柱,杨谦,赵敏.苏云金芽孢杆菌杀虫晶体蛋白基因的研究进展[J].东北林业大学学报,2005,33(1):66-67.
[13] 朱晓阳,闫建平,蔡全信,等.苏云金芽孢杆菌以色列亚种杀蚊幼晶体蛋白的定量分析[A].全国杀虫微生物学术研讨会暨湖北省暨武汉市微生物学会和内蒙古微生物学会[C].武汉:中国微生物学会,2008.
[14] SAMBROOK G L,FRITSCH E F,MANIATIS T,et al.Molecular Cloning[M].Beijing:Science Publishing House,1992.
[15] PIMENTEL D.Environmental and economic effects of reducing pesticide use[J].Bioscience,1991,41:402-409.
[16] 雷萍,张金松,周令,等.苏云金芽孢杆菌以色列亚种对花翅摇蚊作用特性研究[J].微生物学通报,2004(1):17-21.
[17] 刘丽君,张金松,雷萍,等.苏云金芽孢杆菌以色列亚种对摇蚊幼虫毒力的生物测定方法[A].第八次全国环境微生物学术研讨会论文集[C].哈尔滨:中国微生物学会,2006.
[18] 史琦琪,程鹏,王海防,等.苏云金杆菌对微山湖湿地中华按蚊幼虫的室内生测效果[J].中华卫生杀虫药械,2017(3):227-229.
[19] 马素媛,王英,赵清,等.Bti对三带喙库蚊的杀伤效果及温度的影响[J].中国热带医学,2012(12):1427-1429.
[20] 刘刚. 湖南师大“苏云金芽孢杆菌杀虫伴孢晶体蛋白质组学研究”取得重要成果[J].农药市场信息,2009(12):36.
[21] 王利平,代林远,李鹏.苏云金芽孢杆菌研究进展[J].中国畜牧兽医,2011,38(9):224-227.
[22] 关雄. 苏云金芽孢杆菌的研究回顾与展望[J].中国农业科技导报,2006,8(6):5-11.