Stool samples collected during the first days after birth are characterized by low bacterial DNA concentration (12 (link)). For this reason, bacterial DNA was extracted using two different commercial kits to optimize DNA yield. The Power Fecal DNA Isolation Kit (QIAGEN, Valencia, CA) was used for the first stool sample collected according to the manufacturer's instructions, with some modifications, including using a different bead beating tube, the mechanical lysis method, and the incubation temperature. Briefly, a total of 250 μg of stool (wet weight) were placed into a 2 mL Lysing Matrix E tube (MP Biomedicals. Santa Ana, CA). The stool sample and 600 μL of the C1 solution were then heated at 70°C for 10 min. Tubes were then shaken using the FastPrep-24 (MP Biomedicals. Santa Ana, CA) at 6.5 m/s for 45. The remaining steps were performed according to manufacturer's instructions. Bacterial DNA from the subsequent stool samples collected was extracted using the QIAamp Fast DNA Stool Mini Kit (QIAGEN, Valencia, CA), as previously described (3 (link)).
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