绵羊肺炎支原体蛋白质组双向电泳图谱的构建

Establishment of Two-dimensional Gel Electrophoresis pattern in Proteomic of Mycoplasma Ovipneumoniae

为了建立针对绵羊肺炎支原体蛋白质组学研究的双向电泳图谱及其相关技术体系。采用反复冻融兼Urea-CHAPS-DTT-SB3-10裂解法和超声兼Urea-CHAPS-DTT-SB3-10裂解法提取绵羊肺炎支原体菌体蛋白,取200μg菌体蛋白利用固相pH梯度等电聚焦双向电泳,对硝酸银染色后获得的双向电泳图谱进行图像分析。结果表明反复冻融兼Urea-CHAPS-DTT-SB3-10裂解提取法获得198±2个蛋白斑点,超声兼Urea-CHAPS-DTT-SB3-10裂解提取法获得241±2个蛋白斑点。说明超声兼Urea-CHAPS-DTT-SB3-10裂解提取法提高了双向电泳的分辨率,进而可增强生物质谱鉴定蛋白质的准确度。

In order to establish a two-dimensional gel electrophoresis pattern （2-DE） and optimize related techniques in the research of proteomic analysis of mycoplasma ovipneumoniae, two methods （repeating freeze-Urea-CHAPS-DTT-SB3-10 and Sonication -Urea -CHAPS -DTT-SB3-10） were used separately. Electrophorogram were analyzed using bacterial proteins and immobilized pH gradient （IPG） after 2-DE. 2-DE and image analysis revealed 198±9 protein spots were obtained in repeating freeze-Urea-CHAPS-DTT-SB3-10 while 241±11 spots in Sonication-Urea- CHAPS-DTT-SB3-10, which indicated that Sonication-Urea -CHAPS-DTT -SB3-10 improved 2-DE resolving power and increased protein identification accuracy of bio-mass spectrometry.