桶形芋螺毒管蛋白质组双向电泳条件优化

Optimization of Two-dimensional Gel Electrophoresis of Conus betulinus Venom Duct Proteome

目的:优化双向电泳的条件,建立适用于桶形芋螺毒管蛋白质组分析的双向电泳方法。方法:对毒管蛋白的提取、上样量及SDS-PAGE凝胶浓度等影响因素进行优化。结果:乙酸提取法适宜于毒管蛋白的提取,对于pH3～10、17cm的IPG胶条,当上样量为0.75mg,聚焦70000Vhr,SDS-PAGE凝胶浓度为15%时,可提高双向电泳的分离效果,所得蛋白点清晰、数目达到1003个。结论:采用优化的条件进行双向电泳,能得到分辨率高、重现性好、完整的双向电泳图谱,为后续桶形芋螺毒管蛋白质组学研究打下基础。

Objective：To establish a two-dimensional electrophoresis（2-DE） protocol suitable for proteomic analysis of Conus betulinus venom duct.Method：The parameters of the 2-DE were optimized,including protein extraction method,protein loading amount,focusing volt hours,and the gel concentrations for SDS-PAGE etc.Result：The optimal 2-DE conditions were as follows：samples were prepared by acetic acid extraction with pH 3-10 17cm IPG strip,0.75mg loading quantity,70 000Vhr for isoelectric focusing and 15% gel concentrations for SDS-PAGE.Using this protocol,the proteins of the venom duct were well separated,and total 1 003 protein spots were detected.Conclusion：Using 2-DE optimized conditions can get a complete 2-DE map with high resolution and good reproducibility,which would be a strong basis for subsequent research of Conus venom duct proteomics.