表皮葡萄球菌污染对质粒转化和抽提影响的实验研究

Staphylococcus Epidermidis Contamination: A Special Cause of Plasmid Extraction Failure

目的 :探讨导致部分质粒转化和抽提失败的原因。方法 :使用同一批DH5 α细菌制备感受态 ,转化、碱裂解法抽提和鉴定两种质粒 ,经抗性LB琼脂平板筛选和菌落分析 ,对照成功抽提和鉴定的质粒 ,分析部分质粒反复转化和抽提失败的原因。结果 :发现在该抽提失败的质粒所转化的抗性LB琼脂平板上存在两种菌落 ,大量白色菌落为表皮葡萄球菌 ,其过夜振摇产物经碱裂解法抽提质粒失败 ;极少数淡黄色菌落为大肠杆菌 ,经鉴定 ,质粒抽提成功。结论 :隐蔽的表皮葡萄球菌污染可能是导致部分质粒反复抽提失败的重要原因。

Objective: To find the causes of the failure in the extraction of plasmid BAG. Methods:The plasmids of two types (BAG and PET 24 a ) were transformed into the same TSS competent cell DH 5 α Escherichia coli (E. coli). Then under the same conditions, the plasmids were extracted by alkaline lysis and assaying. Screen by the Kanamycin (Kan) selective LB agar plate and analyse the colony. Results:Control plasmid (PET 24a ) can be done exactly well. But the extraction of plasmid BAG failed many times. After transformation, two kinds of colony were observed on the agar plate of BAG, most of the colony is ivory, and few is yellowish; while on the plate of PET 24, only yellowish colony was found. Ivory colony is from Staphylococcus epidermidis and the yellowish is from DH 5 α E. coli. Both bacteria obtained the Kan resistance after plasmid transformation, while Staphylococcus is alkaline lysis resistant, which caused the failure of plasmid extraction. Conclusion: This study suggested that contamination with Staphylococcus epidermidis should be a special cause of transformation and extraction failure to some vectors.