超重力对导入大肠杆菌质粒的诱变效应

The Mutagenic Effect of Hypergravity on Plasmids Conjugated into Escherichia Coli

利用超重力(Hypergravity)处理导入大肠杆菌(DH5α)的质粒,研究超重力对质粒DNA的影响,为超重力作为一种诱变方法提供更充分的理论依据.研究材料经超重力(8500×g,1.5 h)多代定向处理后,利用卡那霉素抗性基因(Kan r)进行筛选,从而验证超重力对基因有一定的诱变效果.在第3代菌株中得到3株突变体,发现突变体与重组菌相比,一条链的序列发生变化.经PCR扩增测序发现:3株突变体突变位点相同,质粒DNA的部分碱基缺失.由于碱基的缺失,产生移码突变,导致提前出现终止子,蛋白质合成终止.对突变菌株继续传代培养得到相同的结果,证明了该突变的可遗传性.通过该研究,初步证实了超重力可以诱发质粒DNA的突变,拓宽了超重力作为一种诱变方法的应用范围.

We use hypergravity to treat the plasmids conjugated into escherichia coli(DH5α)and study Its effect on plasmid DNA with an aim of providing a more sufficient theoretical basis for applying hypergravity as a mutagenesis method.After the multi-generation directional treatment of the research material by hypergravity(8500×g,1.5 h),we screen for mutants in virtue of kanamycin resistance gene(Kan R)in order to verify that hypergravity has a certain mutagenic effect on genes.Three mutants are obtained in the 3 rd generation.Compared with the reorganization bacteria,the configuration of one strand of the mutants have changed.After PCR amplification,we find that the three mutants have the same mutation site and partial base of plasmid DNA is missing.The missing base results in frameshift mutation and the early appearance of terinator and the termination of protein synthesis.Then the third generation of mutant strains are subcultured and the same result is obtained,which proves the heredity of the mutation.This study confirms that hypergravity can induce mutations in plasmid DNA and thus it broadens the application realm of hypergravity as a method for mutagenesis.