Guiney D G, Hasegawa P, Davis C E
Proc Natl Acad Sci U S A. 1984 Nov;81(22):7203-6. doi: 10.1073/pnas.81.22.7203.
A unique shuttle plasmid, pDP1, has been constructed to mediate gene transfer between Escherichia coli and the Gram-negative anaerobe Bacteroides fragilis. pDP1 contains the pBR322 replicon and the Bacteroides clindamycin resistance plasmid pCP1 linked to the transfer origin of the broad host range plasmid RK2. pDP1 can be transferred from E. coli to B. fragilis by the RK2 conjugation system even though RK2 itself is not maintained in the Bacteroides recipients. The antibiotic resistance and replication functions of pDP1 have been mapped by deletion analysis, and a 5-kilobase portion of the plasmid has been identified as the essential region for maintenance in Bacteroides. Comparison of the resistance conferred by pDP1 on E. coli and B. fragilis shows that antibiotic resistance genes are expressed differently in aerobic and anaerobic bacteria. These results document the feasibility of gene transfer from E. coli to B. fragilis and demonstrate the usefulness of this conjugation system to study genetic structure and expression in Bacteroides.
构建了一种独特的穿梭质粒pDP1，用于介导大肠杆菌与革兰氏阴性厌氧菌脆弱拟杆菌之间的基因转移。pDP1包含pBR322复制子和与广宿主范围质粒RK2的转移起始位点相连的脆弱拟杆菌克林霉素抗性质粒pCP1。即使RK2本身在拟杆菌受体中不能维持，pDP1也能通过RK2接合系统从大肠杆菌转移到脆弱拟杆菌。通过缺失分析对pDP1的抗生素抗性和复制功能进行了定位，并且已确定该质粒的一个5千碱基部分是在拟杆菌中维持所必需的区域。比较pDP1赋予大肠杆菌和脆弱拟杆菌的抗性表明，抗生素抗性基因在需氧菌和厌氧菌中的表达不同。这些结果证明了从大肠杆菌向脆弱拟杆菌进行基因转移的可行性，并证明了该接合系统对于研究拟杆菌的遗传结构和表达的有用性。
