RNA-seq and m6A-seq of shIGF2BP2-KYSE30

RNA seq dates of shIGF2BP2 in ESCC cell line KYSE 30. The samples of KYSE30 were subjected to grinding and dissolved in TRIzol (ThermoFisher Scientific, Inc.) for total RNA extraction. Following the evaluation of RNA quality, purity, and integrity, total or poly(A) RNAs were subsequently utilized to generate cDNA libraries. Both transcriptome sequencing and RNA sequencing were conducted using the Illumina HiSeq X Ten platform (Illumina, Inc.). High-throughput transcriptome sequencing was executed at Novogene Co., Ltd. (Beijing, China).m6A-seq dates of shIGF2BP2 in ESCC cell line KYSE 30. Total RNA was obtained and purified from ESCC samples with TRIzolTM (Cat. No. 15596026; ThermoFisher Scientific, Inc.), according to the manufacturer’s instructions. Following extraction, RNA was incubated with m6A antibodies utilizing the Magna Methylated RNA Immunoprecipitation m6A Kit (MilliporeSigma) for immunoprecipitation. The concentration of m6A-modified mRNA underwent examination through m6A-qPCR or m6A-seq methodologies (Haplox Biotechnology Co., Ltd.). Primers targeting the negative region of FOXM1 m6A served as negative controls, whereas primers targeting the positive region of FOXM1 m6A were employed as positive controls. For m6A-seq, the enriched m6A RNA underwent reverse transcription to generate cDNA, and the library was prepared per the instructions provided by Illumina’s NEBNext Ultra RNA Library Preparation Kit (New England Biolabs, Inc.). High-throughput library sequencing was conducted to acquire sequence information about m6A-modified RNA using the Illumina HiSeq XTM Ten platform (Illumina, Inc.).