table S1-S4

This study included 31 patients with papillary thyroid carcinoma (PTC) who were hospitalized in the Third Xiangya Hospital of Central South University from December 2021 to March 2022. Among them, 18 cases were confirmed by postoperative pathology as PTC patients without cervical lymph node metastasis (non-metastatic group), and 13 cases were PTC patients with cervical lymph node metastasis (metastatic group). The exclusion criteria were as follows: patients with other malignant tumors, autoimmune diseases, thyroiditis, and those who had received thyroid-related treatment recently. The clinical information and characteristics of the patients are listed in Table S1. Serum and tissue samples were collected from all patients who underwent thyroidectomy. Total RNA was then extracted using TRIzol reagent (Thermo Fisher Scientific, Massachusetts, USA), and reverse-transcribed into cDNA using an RT kit (Thermo Fisher Scientific, Massachusetts, USA). For the reverse transcription of miRNA and mRNA, Mir-X™ miRNA First Strand Synthesis Kit and mRNA First Strand Synthesis Kit (Thermo Fisher Scientific, Massachusetts, USA) were used respectively to ensure the accuracy and efficiency of the reverse transcription process. Reactions were performed on a real-time fluorescent qPCR system (ABI 7500, New York, USA) using BGreen Premix Ex Taq™ II (TaKaRa, Kyoto, Japan) as the reaction reagent. Parameters such as temperature, time, and number of cycles were strictly controlled during the reaction. The 2-ΔΔCT method was used to evaluate the relative expression of genes, which effectively eliminates experimental errors and accurately reflects the relative expression levels of target genes in different samples. The sequences of the primers used are detailed in Table S2; the primer design was subjected to strict bioinformatics analysis and verification to ensure specificity and amplification efficiency. This study conducted a systematic analysis of the m6A-miRNA regulatory network based on 492 pathologically confirmed PTC patients and 58 normal thyroid tissue samples in The Cancer Genome Atlas (TCGA) database. By integrating transcriptome sequencing and miRNA expression profile data, a co-expression relationship between 30 m6A regulators and 797 miRNAs was constructed (Table S3). Further comparison of expression differences between PTC tissues and normal tissues revealed that among 128 candidate m6A-miRNAs, 47 were significantly upregulated in PTC (e.g., miR-146b-5p, miR-221-3p, etc.), and 36 miRNAs were downregulated (e.g., miR-139-5p, miR-451a, etc.) (screening criteria: |log2 fold change| > 0.40, i.e., fold change > 1.3, and FDR < 0.05) (Table S4).