Ji Xianling, Gai Yingping
College of Forestry, Shandong Agricultural University, Tai'an, Shandong, People's Republic of China.
Methods Mol Biol. 2013;938:339-49. doi: 10.1007/978-1-62703-089-2_29.
Proteome analysis is becoming a powerful tool in the functional characterization of organisms, and takes a broad, comprehensive, systematic approach to understanding biology. Following the sequencing of the phytoplasma genomes, the next step is to characterize the expressed proteome of phytoplasmas to acquire the verification and functional annotation of all predicted genes and their protein products. Here, we describe the protocol of mulberry dwarf phytoplasma purification, phytoplasma protein extraction and separation by SDS-PAGE, in-gel tryptic digestion of the proteins, separation of the digested peptides by liquid chromatography, and identification of the peptides by mass spectrometry. The protocol described here is also applicable to the analysis of other phytoplasma proteomes.
蛋白质组分析正成为生物体功能表征的强大工具,它采用广泛、全面、系统的方法来理解生物学。在植原体基因组测序之后,下一步是表征植原体的表达蛋白质组,以获得所有预测基因及其蛋白质产物的验证和功能注释。在此,我们描述了桑萎缩植原体的纯化、植原体蛋白质提取、通过SDS-PAGE进行分离、蛋白质的胶内胰蛋白酶消化、通过液相色谱分离消化后的肽段以及通过质谱鉴定肽段的实验方案。这里描述的实验方案也适用于其他植原体蛋白质组的分析。