中文摘要：

目的：构建金黄色葡萄球菌（金葡菌）Oat A基因敲除菌株及其回补菌株。方法：以p BT2为载体,构建金葡菌Oat A基因敲除质粒p BT2-△Oat A,经金葡菌RN4220修饰后电转入金葡菌USA300,利用p BT2载体对温度敏感的特点,在42℃多次传代,筛选出金葡菌Oat A基因敲除菌株。以p LI50为载体,构建p LI50-Oat A全基因回补质粒,电转入金葡菌RN4220,再次抽提后电转入敲除菌株△Oat A,获得基因回补菌株p Oat A。结果：成功构建基因敲除质粒p BT2-△Oat A,经RN4220修饰电转入USA300,经PCR及测序鉴定,获得了金葡菌Oat A敲除菌株△Oat A。经PCR及酶切鉴定,确认p LI50-Oat A回补质粒构建成功,通过金葡菌RN4220修饰后电转入敲除菌株△Oat A,获得基因敲除回补菌株p Oat A。结论：成功构建金黄色葡萄球菌Oat A基因敲除菌株及其回补菌株,为进一步研究金葡菌Oat A的功能及其作用机制奠定基础。

英文摘要：

Objective： Construction of OatA deletion mutant and complementation of Staphylococcus aureus（S.aureus）. Methods： Plasmid pBT2-AOatA of S.aureus was constructed on the vector ofpBT2, and used to introduced into S.aureus RN4220 by electroporation. Plasmid pBT2-AOatA of the transformed RN4220, was used to introduced into S.aureus USA300 by electroporation. Taken the advantage of the characteristics of pBT2 vector that was sensitive to temperature, and then repeatedly passaged at 42℃, the OatA deletion mutant S.aureus AOatA was screened. Plasmid pLI50-OatA was constructed on the vector ofpLI50, and used to transformed into S.aureus RN4220 by electroporation. Plasmid pLI50-OatA was purified from the transformed RN4220, and transferred back into S.aureus AOatA. The OatA complementation S.aureus pOatA was obtained. Results： Endonuclease digestion analysis indicated that plasmid pBT2-hOatA was constructed and introduced into S.aureus RN4220 and S.aureus USA300 successfully. The constraction of AOatA S. aureus was confirmed by PCR amplification and sequencing. Endonuclease digestion analysis indicated that plasmid pLI50-OatA was constructed and introduced into S.aureus RN4220 and S.aureus AOatA successfully. The S.aureus pOatA was confirmed by PCR amplification and sequencing. Conclusions： OatA deletion mutant and complementation of Staphylococcus aureus were successfully constructed and would lay the foundation for the further work on the study of OatA function and its mechanism.