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Zhang Y.,China Agricultural University | Zhang Y.,Xinjiang Agricultural Reclamation Academy | Li J.,Xinjiang Agricultural Reclamation Academy | Li J.,Shihezi University | And 2 more authors.
Journal of Animal and Veterinary Advances | Year: 2012

H3K27 methylation plays an important role in the regulation of gene expression and chromatin stability and its expression is dynamically regulated by both specific methylases and demethylases. During development, the regulation of these enzymes is fine-tuned to co-determine the appropriate level of H3K27 methylation thus ensuring correct gene expression. However, it is not completely clear how this regulation is realized. Here, we found that Akt (Aktl andAkt2), H3K27 methylases (Ezhl andEzh2) and demethylases (Jmjd3 and Urx) displayed varying levels of RNA and protein expression in mouse cell lines. Ezh2 was upregulated (p<0.05) at the protein level inanIGF-l-treatedC2C12 cell line. The overexpression of p-Akt through myr-Akt vector transfection greatly downregulated the mRNA levels of Ezhl, Jmjd3 and Utx (p<0.01) but not Ezh2 (p>0.05). Therefore, the data suggest that the expression of both H3K27 methylase and demethylase genes is involved in the activation of the IGF-1/PI3K/Akt pathway. © Medwell Journals, 2012.

Lou Y.-K.,China Agricultural University | Luo J.,China Agricultural University | Dai R.,Xinjiang Agricultural Reclamation Academy | Li N.,China Agricultural University
Progress in Biochemistry and Biophysics | Year: 2010

Myostatin is a TGF-β superfamily member that negatively regulates the growth of the skeletal muscle mass. Remarkable muscle increase was observed in myostatin-knockout mice. Injection and electroporation of myostatin-targeting shRNA into rat tibialis anterior resulted in an increase in its weight, fiber size, and MHC II expression. Two siRNAs targeting mouse myostatin were identified to block mouse myostatin expression upon co-transfection with a myostatin-expressing plasmid into HEK293 cell culture. These siRNAs were cloned into shRNA expression vectors and transferred into C2C12 myoblasts. ShRNA-positive cells were screened by neomycin selection and flow cytometry. By using real-time PCR, it was determined that the endogenous myostatin mRNA expression decreased by 10.2% and 35.5% in Mst-shRNAl-treated and Mst-shRNA2-treated C2C12 myoblasts, respectively. Western blot analysis indicated that the myostatin protein expression level decreased by 29.3% and 64.7%, respectively, in the two groups, it was also demonstrated that downstream MyoD pathway was affected by myostatin blockade, as evidenced by the 24.4% and 40.4% upregulation of MyoD expression in shRNA-treated cells. The results indicate that myostatin-targeting siRNA produced endogenously could efficiently downregulate myostatin expression. This RNAi-based method of increasing muscle mass could provide an alternative strategy to gene knockout methods for genetic breeding and may be useful in improving the economic properties of livestock.

Guan F.,China Jiliang University | Shi G.,Xinjiang Agricultural Reclamation Academy | Wan P.,Xinjiang Agricultural Reclamation Academy | Dai R.,Xinjiang Agricultural Reclamation Academy | And 3 more authors.
Animal Science Papers and Reports | Year: 2014

The FecB gene has been shown to be crucial in reproduction in many sheep breeds. It is a single nucleotide polymorphism (SNP) located in the bone morphogenetic protein receptor IB (BMPRIB) gene. The current methods for genotyping the FecB mutation are either slow and laborious or expensive. In this report, a single-step amplification approach suitable for FecB genotyping method is described. Multiplex PCR was performed with four primers on the basis of tetraprimer amplification refractory mutation system PCR (tetra-primer ARMS PCR), then three FecB genotypes can be detected after electrophoresis. Genotyping results of the proposed multiplex PCR occurred to be in complete accordance with forced PCR-RFLP of all samples. It is a rapid and simple method for detection of FecB in a larger number of samples, and is suitable for other SNPs detection with specific primers even in most "low-tech" laboratories.

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