Supplementary data: Comparison of target enrichment strategies for ancient pathogen DNA

Supplementary Note 1 – Laboratory workflow Supplementary Note 2 - Bioinformatics and Statistical Analysis Supplementary Note 3 – Results of the Bioinformatics and Statistical Analysis Supplementary Figure 1: Comparison of (A) mean coverage, (B) standard deviation of the mean coverage, (C) enrichment factor, (D) and the percentage of the genome covered 5 fold, (E) distribution of the fragment length and (F) frequency of the aDNA damage for the ancient and modern strains of M. leprae. Three independent replicates were performed for each method. Labels of the ancient samples are in black and for the modern samples in red. Boxplots of the array are blue, of the DNA bait capture red and the RNA baits capture is green and grey for the first and second round, respectively Supplementary Figure 2: Comparison of (A) mean coverage, (B) standard deviation of the mean coverage, (C) enrichment factor, (D) and the percentage of the genome covered 5 fold, (E) distribution of the fragment length and (F) frequency of the aDNA damage for the ancient and modern strains of T. pallidum. Three independent replicates were performed for each method. Labels of the ancient samples are in black and for the modern samples in red. Boxplots of the array are blue, of the DNA bait capture red and the RNA baits capture is green and grey for the first and second round, respectively Supplementary Figure 3: Number of unique reads for the three replicate batches of the three tested methods. The number of unique reads in the second round of hybridization with the RNA baits does not strongly increase compared to the first round. Supplementary Table 1: List of all samples used in this study group according to organism and age together with the original publications. Supplementary Table 4: Comparison of the specific reads of the three tested protocols. Supplementary Table 6: Comparison of the variance within each method tested. Supplementary Table 7: Comparison of the costs per reaction.补充说明1——实验室工作流程 补充说明2——生物信息学与统计分析 补充说明3——生物信息学与统计分析结果 补充图1：麻风分枝杆菌（M. leprae）古代与现代菌株的对比分析，包括（A）平均覆盖度、（B）平均覆盖度的标准差、（C）富集因子、（D）基因组5倍覆盖区域占比、（E）片段长度分布，以及（F）古代DNA（aDNA）损伤频率。每种方法均设置3次独立生物学重复。古代样本标注为黑色，现代样本标注为红色。三种检测方法的箱线图分别为：芯片（array）法以蓝色表示，DNA诱饵捕获法以红色表示，RNA诱饵捕获第一轮为绿色、第二轮为灰色 补充图2：梅毒螺旋体（T. pallidum）古代与现代菌株的对比分析，包括（A）平均覆盖度、（B）平均覆盖度的标准差、（C）富集因子、（D）基因组5倍覆盖区域占比、（E）片段长度分布，以及（F）古代DNA（aDNA）损伤频率。每种方法均设置3次独立生物学重复。古代样本标注为黑色，现代样本标注为红色。三种检测方法的箱线图分别为：芯片法以蓝色表示，DNA诱饵捕获法以红色表示，RNA诱饵捕获第一轮为绿色、第二轮为灰色 补充图3：三种受试方法的三次重复批次的唯一读段（unique reads）数量。与第一轮RNA诱饵捕获相比，第二轮的唯一读段数量未出现显著提升 补充表1：本研究所有受试样本按物种及样本年代分组的列表，附原始文献来源 补充表4：三种受试实验方案的特异性读段对比 补充表6：各受试方法内部的方差对比 补充表7：单次反应的实验成本对比