Comparative Proteomics Analysis Of Longissimus Muscle Form Different Breeds Of Cattle

Intramuscular fat content is very important for meat quality. However, the molecular mechanism involved in intramuscular fat deposition is far to completely understand. Qinchuan beef Cattle, Snowdragon Cattle, Yunling Cattle are famous beef cattle breeds in china, of which beef quality is comparable with foreign special beef cattle. It assists in exposure of regulation pathway responsible for intramuscular fat accumulation to compare protein expression of longissimus muscle between beef cattle and Qinchuan cattle.In the current research, Two-Dimensional Gel Electrophoresis and Isobaric Tags for Relative and Absolute Quantitation along with meat analysis, statistical analysis and bioinformatics analysis were used to screen crucial proteins as well as to reveal metabolic pathways related to intramuscular fat accumulation with Chinese primitive Qinchuan cattle and beef cattle as research subjects.1. Intramuscular fat content, tenderness, oleic acid content and water holding capacity of longissimus muscle from beef cattle(Qinchuan beef Cattle, Snowdragon Cattle, Yunling Cattle) are significantly higher than primitive cattle(P<0.05). While the opposite is true for Linoleic acid content(P<0.05). Intramuscular fat content is correlated significantly with tenderness and water holding capacity(P<0.05). More, it also affect unsaturated fattyacid compositions, which is significantly positively related to oleic acid content and palmitoleic acid content(P<0.05), and negatively to Linoleic acid content(P<0.05).2. Two-dimensional electrophoresis technologic platform was established for longissimus muscle with high fat content. Details are showed as follows:(1) The optimum sample preparation method was TCA/acetone precipitation combined with mechanical grinding;(2) Air drying protein precipitate for 30 min was better than 10 min.(3) DTT was a more suitable reducing agent than TBP.(4) 3-10 NL strip had the best separation effect.(5) The most suitable loading sample amount was 200ug;(6) Low room temperature elongated protein separation mode, and changeable room temperature could affect repeatability of 2-DE.(6) The classic silver staining was better than the modified, while the result of the ordinary CBB staining was similar with the colloidal Coomassie staining.(7) Low room temperature affected coloring of silver staining, and the change of room temperature decreaced repeatability of silver staining.(8) We compared and analysed the total protein expression pattern of 4 breeds via the optimized two-dimensional electrophoresis, of which result showed high matching rate of protein spots and detected some differential proteins. However, in view of shortage with two-dimensional electrophoresis, it is necessary to conduct i TRAQ analysis.3. ITRAQ data was processed using discriminant analysis and PCA. Sixteen individuals were classified correctly and four varieties were successfully separated. Variance analysis was used to screen proteins with 1.3 times greater abundance than primitive cattle(P < 0.05). Sixty-seven differential proteins were determined, out of which, 33 proteins were highly expressed in beef cattle and 34 in primitive cattle. The former mainly involved in glycolysis and pentose phosphate pathway. The latter mainly involved electron respiratory chain of transmission, production of energy, muscle contraction and so on. Six proteins remained after combined screening with several statistical methods including fatty acid synthase, fatty acid binding protein, vimentin, annexin, lactate dehydrogenase and NDRG2, which are related to adipocyte proliferation and differentiation(growth of the lipid droplets and fat deposition).