Quantitative reverse transcription‑polymerase chain reaction

YK Yutaka Kawano
MT Maki Tanaka
YS Yasushi Satoh
SS Shigekazu Sugino
JS Jun Suzuki
MF Masaki Fujishima
EO Eri Okumura
HT Hideo Takekoshi
OU Osamu Uehara
SS Shintaro Sugita
YA Yoshihiro Abiko
TT Tetsu Tomonari
HT Hironori Tanaka
HT Hidekatsu Takeda
TT Tetsuji Takayama
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cDNA was synthesized from 500 ng of total RNA using ReverTra Ace quantitative reverse transcription‑polymerase chain reaction (qPCR) RT Master Mix with gDNA Remover (Toyobo, Osaka, Japan) according to the manufacturer's protocol. qRT-PCR was performed in triplicate using KAPA SYBR FAST qPCR Master Mix (Kapa Biosystems, Wilmington, MA, USA) and CFX 96 Connect Real-Time System (Bio-Rad Laboratories Inc., Hercules, CA, USA). Thermocycling conditions were 3 min at 95 °C, followed by 40 cycles of 95 °C for 3 s and 60 °C for 20 s. Changes in relative gene expression between cDNA samples were determined using the 2−ΔΔCq method40. The primers used in this study are listed in Supplementary Table S2.

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