Mirna reverse transcriptase kit

The gastrin (GAS), vasoactive intestinal peptide (VIP), serum motilin (MTL), and cholecystokinin (CCK), Trizol agent and miRNA reverse transcriptase kit were all purchased from Co., Ltd. Trizol agent and miRNA reverse transcriptase kit were both from Invitrogen Company. The EasyPure miRNA kit was from Beijing TransGen Biotech Company, China (ER 601-01). Transscript Green miRNA Two-Step qRT-PCR SuperMix was from Beijing TransGen Biotech Company, China (AQ202-01). KH19A desktop high-speed and high-performance centrifuge was from KAIDA Company. The low-temperature refrigerator (−80°C) was from ThermoFisher Scientific Company. The primer sequence of miR-152 was devised and compounded by Sangon Biotech (Shanghai) Co., Ltd. (Table 1).

The following reagents and instruments were used in this study: dual-luciferase reporter assay system (Promega Madison, US), fetal bovine serum (FBS), Dulbecco's modified eagle medium (DMEM) cell culture medium (Gibco, Rockford, US), radio immunoprecipitation assay (RIPA) buffer (Beyotime, Shanghai, China), Matrigel (BD, New Jersey, US), cell counting kit-8 (CCK8) assay kit (Dojindo Corp, Kyushu, Japan), Gene Mutation Kit and SYBR Green Premix Ex Taq™ II (TaKaRa, Dalian, China), Pierce^TM Magnetic RNA-Protein Pull-Down kit, Lipofectamine 3000, M-MLV reverse transcriptase kit, miRNA reverse transcriptase kit, TRIzol reagent, and SuperSignal West Dura Extended Duration Substrate (Thermo Fisher Scientific, Inc., Waltham, US). Antibodies were purchased from Santa Cruz (Dallas, US). Propidium iodide and APC-Annexin V were purchased form Sigma-Aldrich (St. Louis, USA). The PsiCHECK™-2 vector was purchased from Promega (Madison, US).

RNA from tissues or cells was isolated using Trizol (Vazyme, Nanjing, China). The RNA in nucleus or cytoplasm was extracted with a Cytoplasmic & Nuclear RNA Purification kit (Norgen Biotek, Thorold, Canada). The exosomal RNA was prepared via using Exosome Purification and RNA Isolation kit (AmyJet Scientific, Wuhan, China). circRNA was purified via digesting the linear RNA using RNase R (Geneseed, Guangzhou, China). One μg RNA was used to complementary DNA synthetization using the miRNA Reverse Transcriptase kit or M-MLV Reverse Transcriptase kit (Thermo Fisher Scientific). The complementary DNA was mixed with SYBR (TaKaRa, Otsu, Japan) and specific primers (Sangon) for qRT-PCR. The procedure was set as: 95 °C for 5 min, and 40 cycles of 95 °C for 10 s, 58 °C for 30 s and 72 °C for 1 min. The primer sequences were displayed in Supplementary Table 1. U6 or 18 s rRNA functioned as a normalized reference. Relative RNA level was analyzed via 2^-ΔΔCt method.