RNA isolation and reverse transcription-quantitative real-time PCR (RT-qPCR)
This protocol is extracted from research article:
Overexpression of TC2N is associated with poor prognosis in gastric cancer
J Cancer, Jan 1, 2021; DOI: 10.7150/jca.50653

RNA was isolated from the cell lines using the RNA isolation plus (TaKaRa, Japan). Single-stranded cDNA was generated from 1μg total RNA in a 20μL reaction volume using oligodT primers according to the protocol supplied with the Primer Script TM RT Reagent (TaKaRa, Japan). The relative expression levels were measured by qPCR using the ABI 7900HT instrument (Applied Biosystems, USA) in a total volume of 10ul with the SYBR green detection system (Takara, Japan) and GAPDH was used as an endogenous control. Oligonucleotide sequences of the primer sets used were as follows: TC2N (forward: 5′ TGGCTGTACTGAGGATTATTTGC-3′, reverse: 5′- TGTGAAGGAGTTTCTTGTGTCC-3′); GAPDH (forward: 5′-ACAACTTTGGTATCGTGGAAGG-3′, reverse: 5′-GCCATCACGCCACAGTTTC-3′).

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