Real-time PCR analysis of Slc5a2 expression

Total RNA was extracted from the frozen tissues using TRIzol reagent (Invitrogen) and a quantitative real-time PCR was performed using a StepOnePlus Real-time PCR System and the StepOne Software program (Applied Biosystems), as described previously [28 (link)]. The real-time PCR protocol consisted of one cycle at 95 °C for 20 s followed by 40 cycles at 95 °C for 1 s and 60 °C for 20 s using the primers for Slc5a2 (Mm00453831_m1; Applied Biosystems) and GAPDH (Mn03302249_g1; Applied Biosystems). The transcriptional levels were determined using the ΔΔCt method with normalization to GAPDH.

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Yoshii A., Nagoshi T., Kashiwagi Y., Kimura H., Tanaka Y., Oi Y., Ito K., Yoshino T., Tanaka T.D, & Yoshimura M. (2019). Cardiac ischemia–reperfusion injury under insulin-resistant conditions: SGLT1 but not SGLT2 plays a compensatory protective role in diet-induced obesity. Cardiovascular Diabetology, 18, 85.

Publication 2019

TRIzol reagent StepOnePlus Real-time PCR System StepOne Software program Mn03302249_g1

Frozen Gapdh Primers Real time pcr Slc5a2 Tissues Transcriptional Trizol

Corresponding Organization :

Other organizations : Jikei University School of Medicine

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Variable analysis

independent variables

RNA extraction using TRIzol reagent (Invitrogen)
Real-time PCR using StepOnePlus Real-time PCR System and StepOne Software program (Applied Biosystems)

dependent variables

Transcriptional levels of Slc5a2 and GAPDH genes

control variables

Primers for Slc5a2 (Mm00453831_m1; Applied Biosystems) and GAPDH (Mn03302249_g1; Applied Biosystems)
GAPDH as a reference gene for normalization using the ΔΔCt method

controls

No positive or negative controls were explicitly mentioned in the input protocol.

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