## 2 Materials and Methods
### PCR Amplification and Illumina Sequencing  
PCR amplification of the extracted DNA was performed using primers targeting regions between V3 and V4 of the 165 rRNA gene. For bacterial amplification, 341F primers (5\({}^{\prime}\)TCGTCGGACGCGTC-AGATGTGTATAAGAGACAG-CCTACGGGGNGGCWGGCAG-3\({}^{\prime}\), the target region primer) and 805R (5\({}^{\prime}\)-GTCTGGGCTCGGAGATGGTATAAGAGACA G-GACTACHVGGGATCTAATCC-3) were used. Amplifications were conducted under the following conditions: initial denaturation at 95 \({}^{\circ}\)C for 3 min, followed by 25 cycles of denaturation at 95 \({}^{\circ}\)C for 30 s, primer annealing at 55 \({}^{\circ}\)C for 30 sec, and extension at 72 \({}^{\circ}\)C for 30 s, with a final elongation at 72 \({}^{\circ}\)C for 5 min. Secondary amplification for attaching the Illumina Nextera barcode was then performed using the i5 forward primer (5\({}^{\prime}\)AATGATGGCGCACCAGGATCTACAC-XXXXXXXX-TCGTCGGCAGCGTC-3\({}^{\prime}\); X indicates the barcode region) and i7 reverse primer (5\({}^{\prime}\)-CAAGCAAAGAGAGCCATACG  
\begin{table}
\begin{tabular}{c c c c c c c c} \hline \hline \multirow{2}{*}{**Group**} & \multirow{2}{*}{**Location**} & \multirow{2}{*}{**Maxillary Central**} & \multirow{2}{*}{**Maxillary Right First**} & \multirow{2}{*}{**Maxillary Left First**} & \multirow{2}{*}{**Mandibular Left First**} & \multirow{2}{*}{**Mandibular Left First**} & \multirow{2}{*}{**Mandibular Right First**} & \multirow{2}{*}{**Non-MIM**} \\  & & & **Incisors** & & **Molar** & & **Molar** & **Molar** & **Molar** \\ \cline{3-8} \multirow{4}{*}{**Period**} & \multirow{4}{*}{**Location**} & **11** & **16** & **26** & **36** & **46** & **Nor** \\ \hline \multirow{3}{*}{**Endo**} & **1** & **Supragingival plaque** & 11-1 & 16-1 & 26-1 & 36-1 & 46-1 & Nor-1 \\  & **2** & **Subgingival plaque** & 11-2 & 16-2 & 26-2 & 36-2 & 46-2 & Nor-2 \\  & **3** & **Apical abscess** & & & & & 46-3 & \\ \hline \multirow{3}{*}{**Endo**} & **4** & **Coronal pulp chamber** & 16-4 & * & 36-4 & ** & \\  & **5** & **Root canal** & 16-5 & * & 36-5 & ** & \\ \hline \hline \end{tabular}  
* The crown and root were separated during the extraction; ** Samples were taken but DNA extraction failed due to DNA degradation or insufficient amount.  
\end{table}
Table 1: Sample identification (ID).  
Figure 4: **Sampling of normal and MIM teeth.** Perio group:  
AT-XXXXXXXXAGTCCTGGGGCTCGG-3). The conditions for the secondary amplification were similar to the earlier one except that the amplification cycles were set to 8.  
The PCR products were confirmed using 2% agarose gel electrophoresis and visualized by the Gel Doc system (BioRad, Hercules, CA, USA). The amplified products were purified via the QIAquick PCR purification kit (Qiagen, Valencia, CA, USA). Equal concentrations of purified products were pooled together and short fragments (non-target products) were removed with the Ampure bead kit (Agencourt Bioscience, MA, USA). The quality and size of the products were assessed on a Bioanalyzer 2100 (Agilent, Palo Alto, CA, USA) using a DNA 7500 chip. Mixed amplicons were pooled, and sequencing was conducted at Chunlab, Inc. (Seoul, Korea) using the Illumina MiSeq Sequencing System (Illumina, San Diego, CA, USA) according to the manufacturer's instructions.