Construction of a low-pH-sensing system in Streptococcus mutans

doi:10.7518/hxkq.2017.03.003

Abstract

Abstract:

Objective To construct a low-pH-sensing system in Streptococcus mutans (S. mutans) and to visually detect the pH in situ. Methods Promoter of ureaseⅠ(PureⅠ) and green fluorescence protein (gfp) DNA fragments were amplified by polymerase chain reaction (PCR) from the genome of Streptococcus salivarius 57.I and S. mutans containing the gfp fragment. The two amplified DNA fragments were ligated together and further integrated into pDL278 to construct the recombinant plasmid pDL278-pureⅠ-gfp. This recombinant plasmid was then transformed into S. mutans UA159 cells. Subsequently, the intensity of the optical density per unit area of the low-pH-sensing system was measured and compared under different pH conditions and different processing times. Results PureⅠ and gfp DNA fragments were amplified successfully with the correct molecule sizes (450 and 717 bp, respectively). The recombinant plasmid pDL278-pureⅠ-gfp was constructed and fur-ther verified by PCR and sequencing. The intensity of the optical density per unit area of the low-pH-sensing system increased with decreasing pH and increasing processing time. Conclusion A low-pH-sensing system was constructed successfully in S. mutans. Our research verified that pureⅠ of Streptococcus salivarius can function well in S. mutans as an acid induced promoter, and provided a new method of detecting the pH of plaque biofilms in situ.

Key words: Streptococcus mutans, Streptococcus salivarius, green fluorescence protein, urease, promoter

CLC Number:

R 280.2

Di Kang, Yuqing Li, Xuedong. Zhou. Construction of a low-pH-sensing system in Streptococcus mutans[J]. West China Journal of Stomatology, 2017, 35(3): 239-244.

Figures/Tables 8

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目的基因	引物	产物长度/bp
pureⅠ	pureⅠ-SacⅠ：5’-TTTGAGCTCATTCCTGGGAGACTAGCTG-3’	450
	pureⅠ-BamHⅠ：5’-TTTGGATCCCTCCTAAGTTTTTTATGTTAATATC-3’
gfp	gfp-BamHⅠ：5’-AGAAGGATCCATGAGTAAAGGAGAAGAACTTTTC-3’	717
	gfp-SalⅠ：5’-TTTTGTCGACCTATTTGTATAGTTCATCCATGCCATG-3’