Novaseq600

Paired diagnosis-relapse samples from 6 Pediatric AML patients that were enrolled in AAML1031 were profiled by CITE-seq, including scRNA-seq (Immunai), labeling RNAs with a 10 × Genomics Chromium controller and sequencing with Illumina Novaseq 600. In total, we profiled a total of 15,857 genes in 27,687 cells, with an average of 4,644 UMIs and 1432 gene features per cell (RNA only). Cells with mitochondrial gene content above 10% and fewer than 500 UMIs were excluded. AML samples were treated with RNAlater and profiled using Illumina Novaseq 600 with 25 M reads per sample. Similarly, patients in the NB1 dataset were profiled by bulk RNA-seq with Illumina Novaseq 600 with 25 M reads per sample.

For DNA extraction, the stored JP233 culture was streaked onto LB plate and incubated in a growth chamber at 28 °C for 3 days. A single pure colony was inoculated into a sterile tube containing 5 mL LB broth, which was then grown in an orbital shaker (180 rpm) for 48 h at 28 °C. Strain JP233 was harvested and genomic DNA was extracted using a Wizard^® Genomic DNA Purification Kit (Promega, Madison, WI, USA), following the manufacturer’s instructions.
The JP233 genome was sequenced using a combination of PacBio RS II Single Molecule Real Time (SMRT) and Illumina NovaSeq 600 sequencing platforms by Majorbio Bio-pharm Technology Co., Ltd. (Shanghai, China). For Illumina sequencing, DNA samples were sheared into 400 bp–500 bp fragments and used to build the sequencing library. The prepared library was then used for paired-end Illumina sequencing using the PE150 strategy. For PacBio sequencing, a 10 kb insert library was prepared and sequenced on one SMRT cell using standard methods.

Total DNA extractions were performed on the 200 µL of soil water concentrates, using the DNeasy® PowerSoil® kit (Qiagen). The protocol followed the manufacturer's recommendations: cell lysis, followed by DNA fixation and elution. However, the recommended volume of 100 μL for the elution step was reduced to 60 μL to concentrate the DNA. Final DNA concentrations were quantified using the Invitrogen™ Qubit™ 2.0 fluorometer (Thermo Fischer) and ranged from 0.37 to 1.0 ng µL⁻¹. The extracted DNA was then stored at −20°C until further analyses. The viral communities were determined using shotgun metagenomics sequencing. From the total extracted DNA of soil water samples, library preparations were completed using the Nextera DNA Flex kit (Illumina). Paired‐end sequence reads were generated using the Illumina NovaSeq. 600 (2 × 150 bp).