Longissimus dorsi muscle (LD) proteomics provides a novel opportunity to reveal the molecular mechanism behind intramuscular fat deposition. Unfortunately, the vast amounts of lipids and nucleic acids in this tissue hampered LD proteomics analysis. Trichloroacetic acid (TCA)/acetone precipitation is a widely used method to remove contaminants from protein samples. However, the high speed centrifugation employed in this method produces hard precipitates, which restrict contaminant elimination and protein re-dissolution. To address the problem, the centrifugation precipitates were first grinded with a glass tissue grinder and then washed with 90% acetone (TCA/acetone-G-W) in the present study. According to our result, the treatment for solid precipitate facilitated non-protein contaminant removal and protein re-dissolution, ultimately improving two-dimensional gel electrophoresis (2-DE) analysis. Additionally, we also evaluated the effect of sample drying on 2-DE profile as well as protein yield. It was found that 30 min air-drying did not result in significant protein loss, but reduced horizontal streaking and smearing on 2-DE gel compared to 10 min. In summary, we developed an optimized TCA/acetone precipitation method for protein extraction of LD, in which the modifications improved the effectiveness of TCA/acetone method.

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ABSTRACT. The BM2113 locus was amplified in Yunnan mithun (Bos frontalis) from the southwest mountains of China. It showed a high degree of polymorphism with a total of 12 alleles. The 121-bp polymorphic allele of the BM2113 locus that accounted for 37.1% of homozygotes was the predominant allele with a frequency of 58.57%, identified as mithunspecific for Bos species in Yunnan mithun. The polymorphism information content value was high with a mean of 0.6170, the expected and observed heterozygosity was moderate with values of 0.6427 and 0.6000, respectively, and the BM2113 locus was under Hardy-Weinberg equilibrium (P = 0.2897) in the Yunnan mithun population. This study elucidated the genetic diversity, multi-origin, specific alleles, and characterization of mithun.

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