Weber J M, Johnson S P, Vonstein V, Casadaban M J, Demirjian D C
ThermoGen, Inc., Chicago, IL 60612, USA.
Biotechnology (N Y). 1995 Mar;13(3):271-5. doi: 10.1038/nbt0395-271.
We have developed a chromosomal integration system for gene transfer into the extreme thermophile Thermus flavus. The system relies on integration at the site of leuB (3-isopropylmalate dehydrogenase) which was cloned from T. flavus. The leuB gene was insertionally inactivated in vitro with a thermostable kanamycin-resistance gene and transformed in single-copy into the chromosome of T. flavus on a plasmid vector. Gene replacement strains required leucine for growth, were stably kanamycin-resistant and could grow in the presence of kanamycin at temperatures up to 55 degrees C.
我们开发了一种用于将基因导入嗜热栖热放线菌（Thermus flavus）的染色体整合系统。该系统依赖于在从嗜热栖热放线菌克隆的leuB（3-异丙基苹果酸脱氢酶）位点进行整合。leuB基因在体外被一个耐热卡那霉素抗性基因插入失活，并以单拷贝形式在质粒载体上转化到嗜热栖热放线菌的染色体中。基因替换菌株生长需要亮氨酸，对卡那霉素具有稳定抗性，并且在高达55摄氏度的温度下，在卡那霉素存在的情况下能够生长。
