具有真细菌基因启动子活性的古生菌DNA片段

Chromosome DNA Fragments From Halobacterium halobium Active as Eubacteria Promoters in Escherchia coli

以大肠杆菌启动子探测质粒pKK232-8 为载体,用4 组限制性内切酶分别消化古生菌盐生盐杆菌R1(Halobacterium halobium )的染色体DNA,在体外进行重组,转化E.coliDH-5α感受态细胞,得到12 个转化子(T1～T12),其抗性水平分布在10～500 m g/L的区间内. 将这些转化子所含质粒分别命名为pSX1～pSX12,限制性酶切分析表明,这些质粒各插入了一段不同的外源DNA 片段,杂交分析表明,抗性最强的转化子T11 所含质粒pSX11 上插入了一段来源于盐生盐杆菌R1 染色体的DNA 片段. 该DNA 片段在大肠杆菌中具有启动子功能.

The chromosome DNA of Halobacterium halobium R1 was digesed by BamHⅠ\|SalⅠ、HindⅢ\|SalⅠ、SmaⅠ\|SalⅠ、SmaⅠ\|HindⅢ, and was recombined with a promoter\|probe plasmid pKK232\|8 and transformed into E.coli. The transformants were selected on resistance plates containing ampicillin (Am) and chloramphenicol(Cm).We obtained twelve transformants (T1~T12), the level of Cm resistance fluctuated from 10 mg/L to 500 mg/L. The recombinant plasmids (pSX1~pSX12) carried distinct inserted fragments. Hybridization showed that recombinant plasmid pSX11 carried a inserted fragment from the chromosome of Halobacterium halobium R1. Re\|transformation proved further that the DNA fragment had promoter function in E.coli. Thus, this indicated that DNA fragments from chromosome of archaebacteria( Halobacterium halobium ) may function as eubacteria ( E.coli ) promoters.