产品详情
pECTO金黄色葡萄球菌染色体基因异位互补质粒
基本信息
原核抗性: 氨苄青霉素Amp
克隆菌株: 大肠杆菌DH5α
培养条件: 37℃
质粒简介
pMAD2 derivative designed for chromosomal gene ectopic complementation in Staphylococcus aureu.pECTO a Gram negative/Gram positive shuttle vector derived from pMAD2 designed to integrate DNA sequences between SAOUHSC_00278 and SAOUHSC_00279 (NCTC8325 nomenclature HG003). It carries about 1kb of both sequences. A terminator introduced between the SAOUHSC_00278 and SAOUHSC_00279 sequences prevents polar effects associated with the expression of inserted genes.This a Gram negative/Gram positive shuttle vector with two specialized replicons and antibiotic markers. pBR322 replicon and ampicillin resistance for E. coli. pE194(ts) replicon and erythromycin resistance for S. aureus. In S. aureus, the plasmid propagates at T<30°C, it becomes a suicide vector at T>37°C.
质粒图谱
pECTO金黄色葡萄球菌染色体基因异位互补质粒使用说明:
1、收到质粒干粉后请先5000rpm离心1min,再加入20μl无菌水溶解质粒,室温放置1min;
2、从-80℃冰箱中取出相应的感受态,置于冰盒上解冻,并做好标记;
3、取2μl质粒加至100μl感受态中,冰浴30min;
4、42℃热激90s,再冰浴2min;
5、加入900μl无抗的LB液体培养基,180rpm震荡培养45min;
6、6000rpm离心5min,仅留100ul上清混匀菌体沉淀;
7、混匀后的菌液加至对应抗性的LB平板上,倒入适量玻璃珠,涂匀液体;
8、将平板正向培养1h,再倒置培养12h~16h;
9、挑取单克隆菌落至对应抗性的LB液体培养基中,震荡培养12h~16h,根据实验需要提取质粒。
pECTO金黄色葡萄球菌染色体基因异位互补质粒注意事项:
1、如果您收到的是甘油菌种,请先四区划线,挑取单克隆培养。
2、如果第二天转化平板长的过多,请将质粒按比例稀释后再转化。