Total RNA was extracted using Trizol reagent (Pufei Biotech Co, Ltd, Shanghai, People’s Republic of China) according to the manufacturer’s instructions. Synthesis of cDNA was performed by reverse transcription using Reverse Transcription System reagents (Promega Corporation, Fitchburg, WI, USA). The expression of Syt-7 was quantified by RT-qPCR analysis using SYBR Premix Ex Taq II (Takara, Kyoto, Japan) with GAPDH as an internal reference. The following sequences of primers were used: 5′-TGACTTCAACAGCGACACCCA-3′ (upstream) and 5′-CACCCTGTTGCTGTAGCCAAA-3′ (downstream) for SYT-7 and 5′-ACTCCATCATCGTGAACATCATC-3′ (upstream) and 5′-TCGAAGGCGAAGGACTCATTG-3′ (downstream). Quantitative PCR was performed according to Takara SYBR Master Mix kit instructions: 95°C for 15 s, followed by 45 cycles of 95°C for 5 s and 60°C for 20 s. The relative gene expression levels were calculated and statistically compared using the 2−ΔΔCT analysis program.13 (link)