Analysis Of Difference Expression For CiaH、clpP Gene Of Fluoride-resistant Strain And Parental Strain Of The Streptococcus Mutans

Objective：Dental caries is a bacterial infectious diseases, is a kind of chronic progressive damageof tooth hard tissue, mainly for demineralization of inorganic and organic matterdecompositio.Tooth decay is characterized by high incidence、widely distributed,Is the mostcommon diseases in the mouth.Streptococcus mutans is the main pathogenic bacteriacavities,Its cause caries virulence characteristics mainly include acidogenicity、acidriucity、adhesion.All this time,prevention and treatment of dental caries is a hot issue, The fluoride toprevent tooth decay has been widely recognized in the field of preventive medicine.But,inrecent years by high concentration fluoride long time used in high frequency selectivegrowth of local caused the fluoride resistant strains.The generation of fluoride resistantstrains set a new task.G scholars found that when under the environment of fluoride, thecariogenicity of fluoride-resistant strain was higher than parental strain,Sheng Jiangjuntheorized its acidriucity was stronger than parental strain.But,the mechanism of acidriucityenhancement has not been fully clarified yet.Research has reported a number of acidriucitygenes of streptococcus mutans,including ciaH、clpP.In the preceding work of our projectgroup,we found the dgk、 ffh、 comD genes of fluoride-resistant strain had geneticmutation.All of work through the metheod of constructing the recombinant plasmid、sequencing to determine genetic mutations.The technolpgy of real-time PCR has matured,Quantitative and homogeneous of genetic analysis is the future development trends ofgenetic diagnosis.Selected fluoride-resistant strain and parental strain at different growthstages.Compared the expression quantity,To implement the analysis of expressiondifferences of ciaH、clpP.To reveal the changes of acidriucity mechanism of fluoride-resistant strain and parental strain on the gene expression level,and parsing the reason of acidriucityenhancement,Laying the ground for the research of cariogenic mechanism and recombinantdental caries vaccine of fluoride-resistant strain.Methods：1The activation of streptococcus mutans,fluoride-resistant strains induced in vitro,theidentification of phenotype、physiological and biochemical and16SrDNA.2The determination of growth curve of streptococcus mutans and fluoride-resistantstrain.316sRNA used as a reference gene, designed the primer of ciaH、clpP.4The total RNA of fluoride-resistant strain and parental strain in the logarithmic phaseand stationary phase was extracted,To determine the expression of ciaH、clpP Usingreal-time PCR.Results：1The fluoride-resistant strain of streptococcus mutans has been successfullyconstructed.The identification of phenotype、physiological and biochemical and16SrDNAits still for streptococcus mutans.2Draw the growth curve of fluoride-resistant strain and parental strain;24h isthe logarithmic phase、48h is the stationary phase.3Designed specific primer successfully.4Real-time PCR results showed:(1)At the time of24h、48h,the ciaH gene of fluoride-resistant strain of streptococcusmutans and parental strain,the expression differences were very significant（P<0.01）;Theexpression quantity of fluoride resistant strains is much higher than the parental strain; ciaHgene of fluoride-resistant strain of48h increased obviously than24h.(2)At the time of24h,the clpP gene of fluoride-resistant strain of streptococcus mutansand parental strain,the expression differences were significant(P<0.05);At the time of48h,the expression differences were very significant（P<0.01）;The expression quantity of parental strain of48h is much higher than the24h,The expression quantity offluoride-resistant strain of48h is much lower than the24h,clpP gene of fluoride-resistantstrain of48h significantly reduce than24h.Conclusion:It is significant in fluoride-resistant strain of streptococcus mutans And parental strainof ciaH、clpP gene,It means that the function of fluoride-resistant strain of streptococcusmutans and parental strain has changed, with the relevant acidriucity enhancement.Determine the expreddion differences of significance.The analysis will provide theoreticalbase for the study of specific function of fluoride-resistant strain of ciaH、clpP gene.