Rna easy isolation reagent cat

Total RNA from the collected leaf samples was isolated using RNA-easy^TM Isolation Reagent Vazyme cat (Vazyme, Nanjing, China) according to the manufacturer’s instructions. About 5 μg of RNA was reverse transcribed as single-stranded cDNA with HiScript^®Ⅱ Q RT SuperMix for qPCR (+gDNA wipe) (Vazyme, Nanjing, China). Quantitative real-time RT-PCR (qRT-PCR) was performed in triplicate on the Roche Lightcyler 480II PCR System using the 2× ChamQ Universal SYBR qPCR Master Mix (Vazyme, Nanjing, China). The amplification conditions were 95 °C for 30 s, followed by 40 cycles of 95 °C for 10 s and 60 °C for 30 s. The correlative expression of CsOSCA genes was calculated with the 2^−ΔΔCt method using the housekeeping gene ACTIN as an internal reference [22 (link)]. Statistical analysis was performed using the statistical product and service solution (SPSS) software by one-way ANOVA and Tukey-HSD test, where p < 0.05 indicates significant difference. The gene-specific primers for qRT-PCR are listed in Table S1.

In this study, 12 pairs of ccRCC and para-cancerous tissues were collected from Tongji Hospital of Huazhong University of Science and Technology (Wuhan, China) between January 2022 and May 2022. Total RNA was isolated from ccRCC and normal kidney cells using RNA-easy^TM Isolation Reagent Vazyme Cat (Vazyme, Nanjing, China). A reverse transcription kit (Vazyme) was used to convert the mRNA into cDNA. qRT-PCR was carried out with a SYBR Green kit R701-02 (TransGen Biotech, Beijing, China) in a final volume of 10 mL to detect the expression level of these genes. The data were analyzed using StepOne software (Thermo Fisher Scientific, China). GAPDH was used as a reference gene to calculate the relative expression levels using 2^-DDCT. Primer sequences are listed in the S1 Table.

Total RNA from the mouse cortex was extracted using an RNA-Easy™ Isolation Reagent Vazyme Cat (RC112-01, Vazyme, Nanjing, China). HiScript II Q RT SuperMix for qPCR (+ gDNA wiper) (R223-01,Vazyme, Nanjing, China) was used for reverse transcription. Quantitative PCR was performed using Universal SYBR Green Fast qPCR Mix was acquired from ABclonal (RK21203, Wuhan, China). The fold change of gene expression was calculated using the 2^−ΔΔCt method. The primers were synthesized by tsingke Biotech and presented in Table S1.