Construction of recombinant for generation of Porphyromonas gingivalis minor accessory proteins FimCDE deficient strain

Shanghai Journal of Stomatology ›› 2013, Vol. 22 ›› Issue (6): 623-627.

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Construction of recombinant for generation of Porphyromonas gingivalis minor accessory proteins FimCDE deficient strain

LI Jing-jing ^1,2, GAO Xin ², LI Kuan-yu ², WANG Wen-mei ¹

1.Department of Oral Medicine, School of Stomatology, Nanjing University & Nanjing Stomatological Hospital. Nanjing 210008; 2.Jiangsu Key Laboratory for Molecular Medicine, Medical School of Nanjing University. Nanjing 210093, Jiangsu Province, China

Received:2013-01-14 Revised:2013-05-13 Online:2013-04-12 Published:2013-04-12
Supported by:
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).

Abstract

Abstract: PURPOSE: 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.

Key words: Porphyromonas gingivalis, FimCDE mutant, Recombinant plasmid

CLC Number:

Q782

LI Jing-jing , GAO Xin , LI Kuan-yu , WANG Wen-mei. Construction of recombinant for generation of Porphyromonas gingivalis minor accessory proteins FimCDE deficient strain[J]. Shanghai Journal of Stomatology, 2013, 22(6): 623-627.

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Construction of recombinant for generation of Porphyromonas gingivalis minor accessory proteins FimCDE deficient strain

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