Lertcanawanichakul Monthon
School of Allied Health Sciences and Public Health, Walailak University, Thasala District, Nakhon Si Thammarat, 80160, Thailand.
Curr Microbiol. 2007 Feb;54(2):108-12. doi: 10.1007/s00284-006-0186-3. Epub 2007 Jan 2.
A 6.09-kb plasmids vector pOri253 was constructed from the plasmid pIL253 (5.2 kb) and a 0.89-kb fragment of oriColE1 from pBluescript II KS. The bifunctional plasmid pOri253 conferred erythromycin resistance in both Escherichia coli and Enterococcus faecalis. It has unique sites for EcoRI, BamHI, SalI, and PstI derived from pIL253 and was lost at a low rate in E. faecalis JCM8726 when cultured in Man, Rogosa, & Sharpe broth without antibiotic. The lactococcal promoter P23 was inserted at one end of the pOri253 multicloning site. Gene expression was assessed by an entAI gene, which produced bacteriocin N15. The E. faecalis harboring constructed plasmid carrying P23 (pOrient23) had more antibacterial activity than parental E. faecalis JCM8726 and its clone harboring non-P23-containing plasmid (pOrient), as determined by means of an overlay method.
一个6.09千碱基的质粒载体pOri253由质粒pIL253（5.2千碱基）和来自pBluescript II KS的0.89千碱基的oriColE1片段构建而成。双功能质粒pOri253在大肠杆菌和粪肠球菌中都赋予了红霉素抗性。它具有源自pIL253的EcoRI、BamHI、SalI和PstI的独特酶切位点，并且在不含抗生素的曼、罗戈萨和夏普肉汤中培养时，在粪肠球菌JCM8726中以低速率丢失。乳球菌启动子P23被插入到pOri253多克隆位点的一端。通过产生细菌素N15的entAI基因评估基因表达。通过覆盖法测定，携带构建的含有P23的质粒（pOrient23）的粪肠球菌比亲本粪肠球菌JCM8726及其携带不含P23的质粒（pOrient）的克隆具有更强的抗菌活性。
