plastisphere metagenome (WGS)
Metagenomic sequencing: Metagenomic sequencing was performed on MP biofilms after 210 days of incubation. DNA was extracted using the CTAB method and its quality, concentration, integrity, and purity were assessed using the Agilent 5400 system (Fragment Analyzer 5400; Agilent, Santa Clara, USA). The sequencing library was constructed using the NEBNext UltraTM DNA Library Prep Kit for Illumina (NEB, Ipswich, USA). DNA was fragmented to a size of 350 bp wth the Covaris M220 sonicator (Covaris S2 System; Covaris, Massachusetts, USA). The DNA fragments underwent end polishing, A-tailing, and ligation with Illumina adapters, followed by PCR amplification. After PCR products were purified by the AMPure XP system (Beckman Coulter, Inc., Brea, USA), library quality was assessed on the Agilent 5400 system (Agilent, Santa Clara, USA) and quantified by QPCR (1.5 nM). The qualified libraries were sequenced using the PE150 strategy on the Illumina NovaSeq platform (Illumina, USA), generating high-resolution microbial genomic data. The Raw Data from biofilm samples were obtained by metagenomic sequencing using the Illumina NovaSeq high-throughput sequencing platform. To ensure data reliability, raw sequencing data were preprocessed with Kneaddata (v0.7.4) to remove low-quality reads and contaminants15,16. Taxonomic classification was performed using Kraken2 (v2.0.7-beta) against a custom-built microbial nucleic acid database, derived from the NCBI NT and RefSeq databases. Bracken (v2.0) was subsequently utilized to predict the relative abundance of species in the samples. For functional analysis, qualitycontrolled, and host-filtered sequences were aligned to the UniRef90 protein database using HUMAnN2 (based on DIAMOND)17, generating functional annotations and relative abundance tables across various functional databases.