[Construction of recombinant for generation of Porphyromonas gingivalis minor accessory proteins FimCDE deficient strain].
OBJECTIVE
To construct a recombinant plasmid containing the upstream of fimC and downstream of fimE of Porphyromonas gingivalis, designated as pPHU281-C-Spec-E, which may be further used to knock out fimCDE gene to determine the role of FimCDE in the infection by P. gingivalis.


METHODS
DNA fragments were generated by PCR with the genomic DNA of P. gingivalis strain ATCC 33277 as the template. The upstream fragment of fimC (fragment C) and downstream fragment of fimE (fragment E) were cloned into the suicide plasmid pPHU281 to generate plasmid pPHU281-C-E. The spectinomycin resistance gene was inserted between fragment C and E to construct plasmid Pphu281-C-Spec-E. The recombinant plasmid was verified by restriction enzyme digestion and DNA sequencing.


RESULTS
The recombinant plasmid pPHU281-C-Spec-E was successfully constructed, which was ready for generation of FimCDE-knockout mutant of P. gingivalis.


CONCLUSIONS
The recombinant plasmid pPHU281-C-Spec-E is a tool for construction of FimCDE deficient mutant of P. gingivalis. Supported by National Natural Science Foundation of China (81070839). Team Project of Medical Leaders in Talent and Innovation of Jiangsu Province (LJ201110), Science and Technology Development Plan of Nanjing City(YKK06115) and Medical Science and Technology Development Project of Nanjing City(ZKX1030).
Department of Oral Medicine, School of Stomatology, Nanjing University & Nanjing Stomatological Hospital. Nanjing 210008; Jiangsu Province, China.E-mail:lijingjing900622@163.com.
Jing-jing
JJ
Li
Xin
X
Gao
Kuan-yu
KY
Li
Wen-mei
WM
Wang
D002681:China; D036781:Fimbriae Proteins; D006801:Humans; D010957:Plasmids; D016966:Porphyromonas gingivalis
D036781:Fimbriae Proteins