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Quantitative reverse‐transcription polymerase chain reaction (qRT-PCR)

SJ Shu-fan Ji
SW Sheng-Lian Wen
YS Yu Sun
PH Pi-wei Huang
HW Hao Wu
MH Mao-lin He
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According to the manufacturer’s instructions, TRIzol (TAKARA, Japan) reagent was used to extract the total RNA. cDNA was synthesized using PrimeScript RT Master Mix (TAKARA, Japan). The cDNA combined with Power SYBR Green PCR Master Mix (Thermo Fisher, USA) were used for real-time PCR. The housekeeper gene, GAPDH, was used as a control. After normalizing GAPDH expression, the normal level of STIL expression in five cell lines was expressed as the relative expression and calculated in accordance with the 2-ΔΔCt method. The primer sequences in this study were as follows: GAPDH: 5′-TGACAACTTTGGTATCGTGGAAGG-3′ (F) and 5-AGGCAGGGATGATGTTCTGGAGAG-3′ (R). The following primers were used for STIL: 5′-CCCAACGCCAACTGGAGATTT-3′ (F) and 5-AGTCGGATGGTCTTCTCAGTC-3′ (R). Each experiment was repeated three times.

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