title: WGS of Limosilactobacillus reuteri; isolated from Sigmodon hispidus stool
design: Two frozen stool pellets were taken from a male S. hispidus, weighed, and diluted to 45 mg/mL in sterile 1x PBS. Serial dilutions were plated on Lactobacilli MRS agar (BD Biosciences) and incubated at 37C for 48 h. A colony PCR screen was performed to identify Limosilactobacillus from stool, using species-specific primers [8]. A new PCR reaction was then performed using the universal primers Uni331F/Uni797R [8] targeting 16S rRNA, and purified PCR products were sent for Sanger sequencing. The majority of these colonies were taxonomically classified as Limosilactobacillus reuteri by their 16S rRNA sequence using NCBI BLAST [9]. A single isolate was grown in MRS broth for 48hrs at 37C with no shaking, and genomic DNA was extracted from the bacterial pellet using the DNeasy PowerSoil Kit (Qiagen) using the manufacturers protocol, Genomic library was prepared using the NEBNext Ultra II FS DNA Library Prep Kit for Illumina. Library quality was assessed on an Agilent 2100 Bioanalyzer System using the Agilent High Sensitivity DNA Kit (5067-4626). Library was sequenced with the NovaSeq 6000 (Illumina) at a depth of 40 million reads. FastQC (https://www.bioinformatics.babraham.ac.uk/projects/fastqc/) was used to examine data quality. Trimmomatic [10] was used to remove adaptors and trim low-quality reads. The package ASA3P v 1.3.0 [11] was used for bacterial genome assembly, scaffolding, and annotation following standard protocols. CheckM [12] and BUSCO [13] were used to assess the completeness of assembly and annotation respectively.