Mirna specific taqman mgb probes

The levels of miR-1294 in the 24 OSCC tissues cells and HSC2, HSC4, SAS and KON cells were detected by qRT-PCR. In detail, the total RNA was extracted from the 24 specimens using the TRIzol reagent, according to the manufacturers' s protocol (Invitrogen; Thermo Fisher Scientific, Inc., Waltham, MA, USA). The level of miR-1294 was then detected by TaqMan miRNA RT Real-Time PCR (17 (link)). Single-stranded cDNA was synthesized by the TaqMan MicroRNA Reverse Transcription Kit (Applied Biosystems; Thermo Fisher Scientific, Inc., Waltham, MA, USA), and then amplified by TaqMan Universal PCR Master Mix (Invitrogen; Thermo Fisher Scientific, Inc.) and miRNA-specific TaqMan MGB probes (Applied Biosystems). The U6 snRNA was used for normalization (18 (link)). The c-Myc mRNA levels in the OSCC tissues were assayed by real-time qPCR (delta Cq method of quantification) using the SYBR-Green reagent (Shanghai Shengong Biotechnology Co., Ltd., Shanghai, China) (19 (link),20 (link)). The primers used for c-Myc (human) are listed as follows: F, 5′-AATGAAAAGGCCCCCAAGGTAGTTATCC-3 and R, 5′-GTCGTTTCCGCAACAAGTCCTCTTC-3; GAP DH F, 5′-CTGACTTCAACAGCGACACC-3′ and R, 5′-TAGCCAAATTCGTTGTCATACC-3′. The following thermocycling conditions were maintained: 94°C for 5 min; 30 cycles of 94°C for 45 sec, 55°C for 45 sec and 72°C for 1 min; and 72°C for 10 min (21 (link)).