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A Streptococcus mutans and detection method technology, applied in the field of medical biology, can solve the problems of long time consumption, high homology of conserved regions, difficult antibody preparation process, etc., and achieve accurate detection effect
Inactive Publication Date: 2013-04-24
CHONGQING YUANLUN BIOTECH +1
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① Microbiological detection methods require the isolation of culture medium S. Mutans. There are many kinds of oral bacteria, which usually require selective medium to be isolated and cultivated. Currently, the most commonly used spirobacitracin medium (MSB) cannot effectively isolate S. .Mutans are separated from many bacteria in the oral streptococcal group (such as Streptococcus distalis, Streptococcus sanguis, Streptococcus salivarius, Streptococcus mitis, non-lactinous Streptococcus, Streptococcus hyperkinetic, K. mutans, etc.)
Although the accuracy of the latter is similar to that of PCR, the preparation process of antibodies is very difficult and time-consuming
③Biochip technology fixes a large number of gene probes, gene fragments or antigens (antibodies) on a special carrier (such as a glass slide) in a specific way, and interacts with the sample to be tested under specific conditions. A technique for instrumentation to record, detect, and analyze, which has high sensitivity but expensive instruments and probes and complex operations
Igarashi et al. (Igarashi T, Yamamoto A, Goto N. Microbiol Immunol, 2001, 45 (5): 341-348.) compared the dex Molecular structure discovery: the dex of the four strains all contain an N-terminal signalpeptide, two variable regions and a conserved region, the homology of the conserved region is high, and the homology of the variable region is low
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[0054] The colony on the plate was picked, and the genome was extracted with the "Bacterial Genomic DNA Extraction Kit" (TIANGEN, DP302), and the operation was carried out according to the instructions.
[0055] 3. Double PCR reaction
[0056] Using the DNA prepared by the above method as a template, using dex-F, dex-R, spaP-F and spaP-R as primers respectively, DNApolymerase (TaKaRa Taq TM , Code: DR001AM) for PCR amplification. The reactio...
Embodiment 2
[0059] Example 2. Double-PCR rapid detection of S. Mutans in dental plaque.
[0061] Routine mouthwash with water, rinse with normal saline, sterile cotton balls to prevent moisture, local disinfection with 2.5% iodine tincture, dental probe to collect dental plaque, and immediately put it into an EP tube containing 50 μL of 0.2M NaOH.
[0062] 2. Template DNA preparation
[0063] Vortex the plaque specimen (IKA VORTEX1) for 30s, let it stand at room temperature for 5min, add 50μL of 0.2M HCl / PBS (200mM HCl, 137mM NaCl, 2.7mM KCl, 10mM NaCl 2 HPO 4 , 2mM KH 2 PO 4 ), mix well, centrifuge at 12000g for 5min, and take the supernatant as PCR template.
[0064] 3. Double PCR reaction
[0065] With embodiment 1. see results figure 2 18 dental plaque specimens from caries patients were double positive in PCR detection, and 6 dental plaque specimens from caries-free patients were negative in PCR detection.
[0066]
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technical field [0001] The invention relates to the technical field of medicine and biology, in particular to a double PCR rapid detection method for Streptococcus mutans. Background technique [0002] Dental caries is one of the most common human diseases, with a high incidence rate. WHO ranks it as one of the three major diseases of human beings, along with cancer and cardiovascular diseases. In the latest "Global Oral Health Report" released by the World Health Organization, it is estimated that 5 billion of the world's 6.3 billion people have suffered from dental caries. In developed countries, 60% to 90% of school-age children and most adults have suffered from dental caries. The incidence rate is 50%, and it is as high as 70% to 90% in children. It has become a global problem that threatens human oral health. [0003] Caries is when sugar-containing food (especially sucrose) enters the oral cavity, it is fermented by cariogenic bacteria in the dental plaque to produce...
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