Total RNA was isolated by the Total RNA-extraction kit (Yeasen Biotech, Shanghai, China, #10606ES60) and converted to cDNA by High-Capacity Prime Script RT reagent (Yeasen Biotech, # 11121ES60) according to the appendant instruction. The sequent qPCR analysis was performed by the ABI StepOne TM Real-Time PCR System (Applied Biosystems, Foster City, CA, USA) with SYBR Green Supermix (Takara, Dalian, China, #RR420L). And for detecting the level of miR-760, qRT-PCR was conducted with the using of TaqMan miRNA assays accompanied with specific primer sets (Applied Biosystems).27 (link) The 2-ΔΔCt calculative method was selected for detection the relative change level of each specific gene. The chosen primer sequences (5′-3′) in the qPCR analysis were listed below:
NEAT1-F: 5′-GTTCCGTGCTTCCTCTTCTG-3′,
NEAT1-R: 5′-GTGTCCTCCGACTTTACCAG-3′,
miR-760-F: 5′-TCAATCCACCAGAGCATGGATAT-3′,
miR-760-R: 5′-CTCTACAGCTATATTGCCAGCCA-3′,
TPM1- F: 5′-GGCACCGAAGATGAACTGGACA-3′,
TPM1- R: 5′-GCGTCTGTTCAGAGAAGCTACG-3′,
GAPDH-F: 5′-GTCTCGTCTGACTTCAACAGCG-3′,
GAPDH-R: 5′-ACCACACTGTTGCTGTAGCCAA-3′,
U6–F: 5′-CTCGCTTCGGCAGCACT-3′,
U6-R: 5′-AACGCTTCACTAATTTGCGT-3’.
NEAT1-F: 5′-GTTCCGTGCTTCCTCTTCTG-3′,
NEAT1-R: 5′-GTGTCCTCCGACTTTACCAG-3′,
miR-760-F: 5′-TCAATCCACCAGAGCATGGATAT-3′,
miR-760-R: 5′-CTCTACAGCTATATTGCCAGCCA-3′,
TPM1- F: 5′-GGCACCGAAGATGAACTGGACA-3′,
TPM1- R: 5′-GCGTCTGTTCAGAGAAGCTACG-3′,
GAPDH-F: 5′-GTCTCGTCTGACTTCAACAGCG-3′,
GAPDH-R: 5′-ACCACACTGTTGCTGTAGCCAA-3′,
U6–F: 5′-CTCGCTTCGGCAGCACT-3′,
U6-R: 5′-AACGCTTCACTAATTTGCGT-3’.