Quantitative Gene Expression Analysis in Muscle Tissue

Firstly, the total RNA extraction kit (BioFlux, Beijing, China) was used to extract the total RNA from the muscle (6 replicates per group). All samples were diluted to achieve the same RNA concentration. RNA reverse transcription was carried out using the PrimeScript TMRT reagent kit (Takara, Tokyo, Japan). The primers used in this experiment are shown in Table 4. The qRT-PCR was assayed by SYBR^® Green Master Mix (Toyobo Co., Ltd., Osaka, Japan) with a CFX Connect Real-Time System (Bio-Rad Laboratories, Inc., Hercules, CA, USA), which was previously described by Zhang et al. [33 (link)]. Finally, statistical analysis of all data was carried out with the 2^−ΔΔCT method.