Total RNA was extracted using TRIzol® reagent (Thermo Fisher Scientific, Inc.) according to the manufacturer's instructions from cell samples. From each sample, ~1.5 µg RNA was reverse transcribed into cDNA using a RevertAid First Strand cDNA Synthesis Kit for mRNA detection (Thermo Fisher Scientific, Inc.), or a Mir-X™ miRNA First-Strand Synthesis kit for miRNA detection (Clontech Laboratories, Inc.) according to the manufacturer's instructions. qPCR was performed using a NovoScript® SYBR Two-Step RT-qPCR kit (Novoprotein Scientific, Inc.) on a QuantStudio™ 6 Flex RT-qPCR system (Applied Biosystems; Thermo Fisher Scientific, Inc.). Thermocycling conditions were initial hold at 95°C for 10 min, 40 cycles of 95°C for 15 sec and 60°C for 60 sec. The sequences of the primer pairs used for qPCR are listed in Table SI. GAPDH and U6 were used as the internal controls. The relative expression of each target was analyzed using the 2−ΔΔCq method (29).

Note: The content above has been extracted from a research article, so it may not display correctly.

Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.

We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.