Study on Construction of Homologous Recombination Plasmid of sigB in Staphylococcus Aureus
- VernacularTitle:金黄色葡萄球菌sigB基因同源重组质粒构建的研究
- Author:
Fengjun SUN
;
Peiyuan XIA
- Publication Type:Journal Article
- Keywords:
Staphylococcus aureus;
SigB gene;
Homologous recombination;
PCR
- From:
China Pharmacy
2001;0(08):-
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:To construct the homologous recombination plasmid of sigB gene from staphylococcus aureus so as to provide the basis for further study of the formation mechanism of its biofilm.METHODS:The target fragment“upstream sequence of sigB,kana-resistance gene,and downstream sequence of sigB”were amplified by PCR from staphylococcus aureus DNA and pEGFP-N2plasmid,respectively,in which the terminals of3interlinked fragments were complementary,and re-stricted enzyme EcoRⅠand XhoⅠsite existed in the sequence terminal,respectively.The fragments were ligated with fusion PCR in turn.Then target fragment was recombinated into plasmid pBluescriptⅡSK(+)according to the instruction manual of commercial kits.After both recombination of pBluescriptⅡSK(+)and vector plasmid pGEM-7zf(+),restricted enzyme SacⅠand XhoⅠ,the homologous recombination plasmid pGEM-7zf(+)-sigB was finally made via T 4 ligation enzyme.RE-SULTS:The target fragments were amplified by PCR and were confirmed with the agarose gel electrophoresis and sequencing.CONCLUSION:The homologous recombination plasmid of sigB gene from staphylococcus aureus is constructed successfully.
