Rna trizol reagent

To validate the expression levels of m⁶A-related prognostic lncRNAs in tumor samples and normal tissues, we performed quantitative real-time polymerase chain reaction (qRT-PCR) assay. Total RNA were isolated from 18 samples using RNA TRIzol reagent (Tiangen Biotech Co., Ltd., Beijing, China, #DP451). Then, cDNA synthesis was performed by using PrimeScript™ RT Master Mix (Takara Biomedical Technology Co., Ltd., Beijing, China, #RR036Q). Real-time PCR was conducted with TB Green Premix Ex Taq (Takara Biomedical Technology Co., Ltd., Beijing, China, #RR820A). Relative expression of lncRNAs was normalized to GAPDH and calculated by using 2-ΔΔCt method. Primer sequences used in our study are as follows: RPP38-DT-F: 5′-CCATCGGAGTCGCTGCAAAGTC-3′, RPP38-DT-R: 5′-AGGAGGAGGCTCATTAGGTCAGAAG-3′; AC024270.4-F: 5′-TCATGAGCCACGAAGTCAAGC-3′, AC024270.4-R: 5′-AGCCTTAAGTCTCAGGTCCTC-3′; AC008124.1-F: 5′-TGCCAACGACTTCTACCACCT-3′, AC008124.1-R: 5′-AGTCACCTCAGCTTTCCGTTC-3′; and AC025176.1-F: 5′-CTTCAACTGGCTTCCTTGCTT-3′, AC025176.1-R: 5′-ACAGGAAACTCCTTCGTCACA-3′.

Total RNA was isolated from 12 samples using RNA TRIzol reagent (Tiangen Biotech Co., Ltd., Beijing, China, #DP451). cDNA synthesis was conducted with PrimeScriptTM RT Master Mix (Takara Biomedical Technology Co., Ltd., Beijing, China, #RR036Q). Real-time PCR was then performed with TB Green Premix Ex Taq (Takara Biomedical Technology Co., Ltd., Beijing, China, #RR820A). Relative expression of lncRNAs were normalized to GAPDH and calculated by 2-ΔΔCt method. Primers sequences are listed in Supplementary Table 2.