Computational protocol: Mitofusion-2-mediated alleviation of insulin resistance in rats through reduction in lipid intermediate accumulation in skeletal muscle

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Protocol publication

[…] Total RNA was extracted from frozen skeletal muscle using a standard TRIzol (Invitrogen, USA) RNA isolation method. Reverse transcription of RNA was carried out according to the instructions of the Easy Script First-Strand cDNA Synthesis Super Mix kit (Trans Gen Biotech, CA). Specific primers designed for amplification of MFN2, CPT1, CD36 and β-actin were verified by NCBI Blast; primers sequences are shown in Table . Real-time PCR was performed on an ABI PRISM 7300 PCR System (Applied Bio systems, USA) using SYBR Green I GoTaq® qPCR Master Mix (Promega, USA). PCRs were carried out in a total of 25 μl as follows: one cycle at 95°C for 5 min, followed by 40 cycles of 95°C for 15 s, 58°C for 20 s and 72°C for 30 s. The gene expression from each sample was analysed in duplicates and normalized against β-actin. The results are expressed as relative gene expression using the ΔCt method. […]

Pipeline specifications

Software tools MUSCLE, BLASTN
Application Nucleotide sequence alignment
Organisms Rattus norvegicus
Diseases Diabetes Mellitus, Muscular Diseases