刺糖多孢菌电转化条件研究

中国生物工程杂志

2011, Vol. 31

Issue (5): 81-85

研究报告

刺糖多孢菌电转化条件研究

张求学^1,2, 兰周², 汪洋², 王昌禄¹, 宋渊³, 张晓琳²

1. 天津科技大学食品工程与生物技术学院天津 300457;
2. 国家粮食局科学研究院北京 100037;
3. 中国农业大学生物学院北京 100094

Study on the Conditions of Electrotransformation in Saccharopolyspora spinosa

ZHANG Qiu-xue^1,2, LAN Zhou², WANG Yang², WANG Chang-lu¹, SONG Yuan³, ZHANG Xiao-lin²

1. College of Food Engineering and Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, China;
2. Academy of State Administration of Grain, Beijing 100037, China;
3. College of Biological Sciences, China Agricultural University, Beijing 100094,China

全文: PDF(1078 KB) HTML

摘要：

以经理化诱变选育的多杀菌素高产菌株刺糖多孢菌(Saccharopolyspora spinosa)CB11为受体菌,将具有安普霉素(apramycin)抗性标记的整合型载体pSET152作为质粒供体,研究了制备刺糖多孢菌感受态细胞时菌体的生长阶段、质粒DNA浓度、电场强度等因素对电转化效率的影响,结果表明,在菌体培养48h至对数中期制备感受态,电场强度为12kV/cm,最小DNA浓度为0.1μg时可获得最高的转化效率,转化子的安普霉素抗性基因的PCR扩增及抗性稳定性实验从分子水平及细胞水平验证了pSET152质粒能整合在刺糖多孢菌的基因组中并能稳定遗传,所有结果表明所建立的刺糖多孢菌电转化体系是行之有效的。

关键词： 刺糖多孢菌; 电转化; 质粒; pSET152

Abstract:

Saccharopolyspora spinosa CB11was used as the recipient strain . Optimal conditions including growth stage of the strain, electroshock voltage,DNA concentration were investigated for the electrotransformation of CB11 with Escherichia coli-Streptomyces shuttle vector pSET152.It was showed that the highest electroporation efficiency was yielded under the cluture age of 48h, electric field strength of 12kV/cm and minimum DNA concentration needed 0.1μg . Plasmid stability experiment and PCR of acc(3)IV gene showed that pSET152 was successfully electroporated into and could stably exist in the Saccharopolyspora spinosa CB11. This protocol would be useful for genetic studies of S. spinosa.

Key words: Saccharopolyspora spinosa Electroporation Plasmid pSET152

收稿日期: 2011-01-20 出版日期: 2011-05-27

ZTFLH:

Q78

基金资助:

"十一五" 国家科技支撑计划资助项目(2006BAD08B02-1)

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引用本文:

张求学, 兰周, 汪洋, 王昌禄, 宋渊, 张晓琳. 刺糖多孢菌电转化条件研究[J]. 中国生物工程杂志, 2011, 31(5): 81-85.

ZHANG Qiu-xue, LAN Zhou, WANG Yang, WANG Chang-lu, SONG Yuan, ZHANG Xiao-lin. Study on the Conditions of Electrotransformation in Saccharopolyspora spinosa. China Biotechnology, 2011, 31(5): 81-85.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/ 或 https://manu60.magtech.com.cn/biotech/CN/Y2011/V31/I5/81

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