Effect of Saliva Collection Methods on the Detection of Periodontium-Related Genetic and Epigenetic Biomarkers—A Pilot Study
Abstract
:1. Introduction
2. Results
2.1. Periodontal Status of Participants
2.2. Salivary Flow Rate, DNA/RNA/Protein Quantity and Quality
2.3. Osteogenic and Wnt Pathway-Related Gene Expression
2.4. DNA and RNA Methylation Epigenetic Factors Changes
2.5. Four Periodontium-Associated DNA Methylation Levels in Saliva from Different Collection Methods
2.6. Correlation between Gene Methylation Levels and Clinical Parameter
3. Discussion
4. Materials and Methods
4.1. Study Participant and Saliva Sample Collection
4.2. Salivary RNA Extraction Using a Trizol™ Method
4.3. Salivary gDNA Isolation Using Trizol
4.4. Salivary Protein Isolation and Quantification
4.5. gDNA Real-Time Quantitative PCR (qPCR)
4.6. mRNA Expression for Osteogenic, Wnt-Related Genes and DNA/RNA Methylation Regulators
4.7. Bisulfite Conversion and Quantitative Methylation-Specific PCR (qMSP)
4.8. Statistical Analysis
5. Conclusions
Author Contributions
Funding
Conflicts of Interest
References
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Gender | Age | PPD | BOP | Collection Method | Ethnicity | |
---|---|---|---|---|---|---|
1 | F | 33 | <4 mm | 27% | Spitting, drooling | Asian |
2 | M | 29 | <4 mm | 41% | Spitting, drooling | Asian |
3 | M | 38 | <4 mm | 14% | Spitting, drooling | Caucasian |
4 | F | 38 | <4 mm | 10% | Spitting, drooling | Asian |
5 | M | 24 | <4 mm | 6% | Spitting, drooling | Caucasian |
Spitting | Drooling | |
---|---|---|
1 | 9 | 4 |
2 | 9 | 2 |
3 | 9 | 3 |
4 | 9 | 1 |
5 | 9 | 1 |
Gene | Forward (5′-3′) | Reverse (5′-3′) | Product (bp) | |
---|---|---|---|---|
RUNX2 | M pair | AGATTTCGTTCGGTAGTCGG | CTCACGTCGCTCATTTTACC | 257 |
U pair | AGATTTTGTTTGGTAGTTGG | CTCACATCACTCATTTTACC | 257 | |
CEMP1 | M pair | TTACGAGGTGTAGAGGTTCGG | ACTCTCAAAACTAATAAAAATAACCCGT | 200 |
U pair | TTATGAGGTGTAGAGGTTTGGA | ACTCTCAAAACTAATAAAAATAACCCATTT | 200 | |
IL 6 | M pair | GAGTTTATCGGGAACGAAAG | CTCCCTCACACAAAACTCGAC | 133 |
U pair | GAGTTTATTGGGAATGAAAG | CCCTCACACAAAACTCAACC | 131 | |
TNF α | M pair | GCGATGGAGAAGAAATCGAG | AACAACTACCTTTATATATCCCTAAAACG | 153 |
U pair | TGATGGAGAAGAAATTGAGAT | AACTACCTTTATATATCCCTAAAACAAAAA | 149 |
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Han, P.; Ivanovski, S. Effect of Saliva Collection Methods on the Detection of Periodontium-Related Genetic and Epigenetic Biomarkers—A Pilot Study. Int. J. Mol. Sci. 2019, 20, 4729. https://doi.org/10.3390/ijms20194729
Han P, Ivanovski S. Effect of Saliva Collection Methods on the Detection of Periodontium-Related Genetic and Epigenetic Biomarkers—A Pilot Study. International Journal of Molecular Sciences. 2019; 20(19):4729. https://doi.org/10.3390/ijms20194729
Chicago/Turabian StyleHan, Pingping, and Sašo Ivanovski. 2019. "Effect of Saliva Collection Methods on the Detection of Periodontium-Related Genetic and Epigenetic Biomarkers—A Pilot Study" International Journal of Molecular Sciences 20, no. 19: 4729. https://doi.org/10.3390/ijms20194729
APA StyleHan, P., & Ivanovski, S. (2019). Effect of Saliva Collection Methods on the Detection of Periodontium-Related Genetic and Epigenetic Biomarkers—A Pilot Study. International Journal of Molecular Sciences, 20(19), 4729. https://doi.org/10.3390/ijms20194729