Bicinchoninic acid (bca)

An equivalent amount of Inner Mongolia Cashmere goat skin samples were ground in liquid nitrogen, followed by the addition of 1% sodium dodecyl sulfate(SDS)to the lysate. The mixture was incubated at room temperature for 20 min, followed by ultrasonication for 2 min and centrifugation at 12,000 rpm for 15 min at 4 °C. The total protein concentration in the supernatant was determined using the BCA (Bioteke,Bei Jing, China) kit. The protein was hydrolyzed by trypsin, and peptides were analyzed by Triple TOF 5600 (AB Sciex, USA). The protein candidates were searched by software Protein Pilot 4.0. The sequences of vimentin in Capra hircus, Ovis aries, Bos taurus, Homo sapiens, and Mus musculus were obtained from NCBI/Protein and compared to that of NCBI/BLAST. The three-dimensional structure of vimentin was constructed by the comparative protein modeling program SWISS-MODEL.

RIPA (Beyotime, China) were used to extract proteins from H9c2 cells. Protein concentrations were determined using BCA (BioTeke, China). After separation on SDS-PAGE, proteins were transferred to a polyvinylidene fluoride (PDVF) membrane. The membrane was then treated with 5% skimmed milk for 1 h. Then, the membrane was incubated with anti-TNRC6A (1 : 500, ABCAm, USA), anticleaved caspase-3 (1 : 500, ABCAm, USA), anti-BCL2 (1 : 1,000, ABCAm, USA), anti-Bax (1 : 1,000, ABCAm, USA), anti-cyclinD1 (1 : 1,000, ABCAm, USA), and GAPDH anti-cyclinD1 (1 : 1,000, ABCAm, USA) at 4°C overnight, followed by further incubation with goat anti-rabbit IgG H and L-HRP (1 : 5,000, ABCAm, USA). SuperECL (Applygen, China) was used to detected membrane-bound immune complexes. GAPDH was used as an internal control.