橡胶树死皮病胶乳C-乳清差异表达蛋白质的筛选与鉴定

Screening and Identification of Differential Expressed Proteins in C-serum on Latex of Tapping Panel Dryness (TPD) in Rubber Tree (Hevea brasiliensis Muell. Arg.)

橡胶树死皮病(tapping panel dryness,TPD)在世界各橡胶种植园普遍发生,给橡胶种植业带来严重的危害。为了更好地了解和阐明死皮病发生、发展的分子机制,应用双向凝胶电泳技术(2-DE)比较橡胶树死皮株与健康株胶乳C-乳清蛋白质组表达的差异。采用固相pH梯度双向凝胶电泳分离橡胶树死皮株与健康株C-乳清的总蛋白质,凝胶经考染显色后,用PDQuest7.40图像分析软件进行比较分析,识别差异表达的蛋白质。这些点经过胶内酶切后进行基质辅助激光解析电离飞行时间质谱(MALDI-TOF-MS)分析获取肽质指纹图谱(PMF),Mascot软件搜索SWISS-PROT和NCBInr数据库鉴定蛋白质。结果:(1)橡胶树死皮株与健康株C-乳清凝胶的平均蛋白质点数分别为1075±35和1134±27,其平均匹配的点数分别为982±38和1008±22,组内图像匹配率达91.89%和88.72%。(2)橡胶树死皮株与健康株C-乳清组间的平均匹配蛋白点数为970±25。利用MALDI-TOF-MS质谱技术对40个差异明显的蛋白点进行分析鉴定,通过查询数据库鉴定了27个蛋白质。建立了分辨率高且重复性较好的橡胶树死皮株与健康株胶乳C-乳清的双向凝胶电泳图谱,并应用质谱技术鉴定了其中表达差异的蛋白质点,这些差异表达的蛋白质可能参与了死皮发生和发展的过程。

Tapping panel dryness （TPD）, which is characterized by the partial or complete cessation of latex flow upon tapping, widely occurs in most of high latex yielding rubber （ Hevea brasiliensis） plantation around the world and leads to severe loss to the natural rubber production. For better understanding and elaborating the mechanism of Tapping Panel Dryness （TPD） of Hevea brasiliensis onset, differential proteomic analysis is conducted in C-serum proteins from latex of the healthy and TPD trees by two-dimension gel electrophoresis （2- DE） . The C-serum proteins from latex of healthy and TPD trees were separated by immobilized （IPG） based 2-DE. The Commassie brilliant blue-stained 2-DE were scanned with digital Image pH gradient Scanner and analyzed with PDQuest7. d0 analysis software to identify the differential proteomic. After in-gel protein digestion, peptide mass fingerprint （PMF） of different-expressed protein spots was obtained by virtue of matrix-assisted laser desorption ionization time-of-flight mass spectrometry （MALDI-TOF-MS）, proteins were identified in Swiss-Prot and NCBInr database by Mascot software. The results of above are as follows ： （ 1 ） Average protein spots were 1075±35, 1134 ± 27 in C-serum proteins from latex of the healthy and TPD respectively, and the average matched spots were 982 ± 38,1008± 22 in both the two types of tissues respectively , and the matching rate was 91.89 %,88.72 % respectively. （2）A total of 970±25 protein spots was matched between the 2-DE map of C- serum proteins from latex of the healthy and TPD. 40 spots at the remarkable level of changed expression were measured by MALDI-TOF-MS and 27 proteins were identified through database searching. The well-resolved and reproducible 2-DE patterns of C-serum proteome from latex of the healthy and TPD trees were established and some different-expressed protein spots between the two types of tissues were detected by mass spectrometry. These proteins may be involved the onset of TPD syndrome. The results presented here demonstrate that 2-DE technique provides a powerful complementary approach for the identification of TPD related proteins from rubber tree.